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1040-9238/02/$.50© 2002 by CRC Press LLC

Critical Reviews in Biochemistry and Molecular Biology, 37(2):71–119 (2002)

Essentiality of Mitochondrial OxidativeMetabolism for Photosynthesis:Optimization of Carbon Assimilation andProtection Against Photoinhibition

K. Padmasree, L. Padmavathi, and A.S. Raghavendra*

Department of Plant Sciences, School of Life Sciences, University of Hyderabad,Hyderabad - 500 046, India

Referee: Christine Foyer, Dept. of Biochemistry and Physiology, IACR - Rothamsted,Harpemden, Herts AL5 2JQ, England, United Kingdom

* Author for correspondence.

** Name and address of corresponding author: Professor A.S. Raghavendra, Department of Plant Sciences, Schoolof Life Sciences, University of Hyderabad, Hyderabad 500 046, India. Tel: +91-40-3010630. Fax: +91-40-3010145 E-mail: asrsl@uohyd.ernet.in

Table of Contents

I. Introduction to the Topic .................................................... 73A. Scope of the Present Review ........................................... 74B. Differential Effects on CO2 Efflux/O2 Uptake ................. 74C. Light Enhanced Dark Respiration (LEDR)...................... 75D. Mitochondrial Respiration in Light: Modified TCA

Cycle ................................................................................. 78

II. Essentiality of Mitochondrial Respiration forPhotosynthesis...................................................................... 79A. Restriction of CO2 Assimilation, but Not of Photochemical

Activities ........................................................................... 81B. Importance at Both Limiting and Optimal CO2............... 82C. Role of Cytochrome and Alternative Pathways of

Oxidative Electron Transport ........................................... 85D. Pronounced Interaction in Algal Mutants/Guard Cells ... 86

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III. Protection Against Photoinhibition .................................... 87A. Oxidative Electron Transport and Oxidative

Phosphorylation ................................................................ 88B. Sustenance of Repair Mechanism by Mitochondrial

ATP ................................................................................... 89C. Photorespiratory Reactions ............................................... 89D. Significance Under Temperature or Water Stress ............ 91

IV. Optimization of Photosynthetic Carbon Assimilation..... 92A. Sustenance of Sucrose Biosynthesis: Role of ATP ......... 93B. Maintenance of Redox state: Ratios of malate/OAA

and triose-P/PGA.............................................................. 95C. Shortening of Induction .................................................... 97D. Activation of Enzymes ..................................................... 99E. Integration with Photorespiration and Nitrogen

Metabolism ..................................................................... 100F. Role in C4 Photosynthesis .............................................. 101

V. Biochemical Basis: Interorganelle Interaction............... 104A. Major Products of Organelle Metabolism ..................... 104B. Metabolite Exchange between Chloroplasts, Mitochondria,

Peroxisomes, and Cytosol .............................................. 106

VI. Future Perspectives........................................................... 108

ABSTRACT: The review emphasizes the essentiality of mitochondrial oxidative metabo-lism for photosynthetic carbon assimilation. Photosynthetic activity in chloroplasts andoxidative metabolism in mitochondria interact with each other and stimulate their activities.During light, the partially modified TCA cycle supplies oxoglutarate to cytosol and chlo-roplasts. The marked stimulation of O2 uptake after few minutes of photosynthetic activity,termed as light enhanced dark respiration (LEDR), is now a well-known phenomenon. Boththe cytochrome and alternative pathways of mitochondrial electron transport are importantin such interactions. The function of chloroplast is optimized by the complementary natureof mitochondrial metabolism in multiple ways: facilitation of export of excess reducedequivalents from chloroplasts, shortening of photosynthetic induction, maintenance ofphotorespiratory activity, and supply of ATP for sucrose biosynthesis as well as othercytosolic needs. Further, the mitochondrial oxidative electron transport and phosphoryla-tion also protects chloroplasts against photoinhibition. Besides mitochondrial respiration,reducing equivalents (and ATP) are used for other metabolic phenomena, such as sulfur ornitrogen metabolism and photorespiration. These reactions often involve peroxisomes and

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cytosol. The beneficial interaction between chloroplasts and mitochondria therefore extendsinvariably to also peroxisomes and cytosol. While the interorganelle exchange of metabo-lites is the known basis of such interaction, further experiments are warranted to identifyother biochemical signals between them. The uses of techniques such as on-line massspectrometric measurement, novel mutants/transgenics, and variability in metabolism bygrowth conditions hold a high promise to help the plant biologist to understand thisinteresting topic.

KEY WORDS: alternative pathway, chloroplasts, cytochrome pathway, interorganelleinteraction, mitochondria, peroxisomes.

I. INTRODUCTION TO THETOPIC

Photosynthesis, the primary source ofenergy for the living world, consists of twodistinct phases. The first phase of photo-chemical reactions, involves the conversionof radiant solar energy into chemical formslike ATP and NADPH (or reduced ferre-doxin) with concomitant evolution of oxy-gen. In the second biochemical phase, theATP and NADPH (or reduced ferredoxin)are utilized to reduce carbon dioxide (orother compounds like NO2 or SO2) intoenergy rich carbon (or nitrogen or sulfur)compounds. Respiration on the other handinvolves the oxidation of carbon compoundsand production of NADH or FADH withthe simultaneous release of CO2. The reduc-tants (NADH or FADH) are oxidizedthrough the electron transport (and oxida-tive phosphorylation) to produce ATP, in-volving consumption of O2 and release ofwater.

Thus, photosynthesis is a process ofreduction and respiration is a process ofoxidation. Both processes provide ATP forcellular needs. The nature of these twometabolic pathways implies that theycomplement each other. The major sites ofphotosynthesis and respiration are chloro-plasts and mitochondria. Although chloro-plasts and mitochondria are traditionally

considered to be autonomous organelles,recent literature has established that thesetwo organelles are not only interdependentin their functions but also are mutually ben-eficial in their interaction.

Besides the carbon metabolism, the re-duced equivalents are consumed in meta-bolic reactions of photorespiration, nitrateassimilation, and sulfur metabolism. Forexample, the requirement of NADH forhydroxypyruvate reduction in peroxisomesis met by chloroplasts as well as mitochon-dria. Naturally, the cytoplasm is a commonmedium for the flux of all related metabo-lites. Thus the interaction of chloroplastsand mitochondria is not exclusive but ex-tends to cytoplasm and peroxisomes.

Under limiting CO2, photorespiration ishighly active and becomes a major linkbetween chloroplasts, peroxisomes, cyto-plasm, and mitochondria. Glycine is themajor substrate of mitochondrial respira-tion under limiting CO2 and can contributesignificant amounts of ATP to cell. At highCO2, the enhanced requirement of ATP incytosol (for sustenance of sucrose biosyn-thesis) is met again from mitochondria(which can use both glycine and malate asrespiratory substrates). Under both situa-tions, nitrogen metabolism and recycling ofammonia/keto acids are always integratedwith the functioning of chloroplasts, mito-chondria, peroxisomes, and cytoplasm. Anymodulation of respiration leads to changes

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in the patterns of photosynthesis and photo-respiration and subsequently modificationof nitrogen as well as sulfur metabolism.

The current review attempts to criticallyassess and emphasize the physiological andbiochemical features of interorganelle inter-action: chloroplasts, mitochondria, peroxi-somes, and cytoplasm. However, emphasisis given to the essentiality of mitochondrialrespiration for photosynthetic carbon metabo-lism. The mitochondrial oxidative metabo-lism not only helps to optimize photosynthe-sis in varied environmental conditions butalso protects the chloroplasts against thephotoinhibition of photosynthesis.

A. Scope of the Present Review

Due to the intriguing but interestingnature of the topic, considerable effort hasbeen made in the last decade to study theoccurrence of mitochondrial respiration inlight and its importance for photosynthesis,particularly in mesophyll protoplasts andleaves. In view of the limited space, all theoriginal articles are not referred to in thisreview. Readers interested in the extensiveliterature in this and related areas may con-sult the previous reviews (Azcón-Bieto,1992; Raghavendra et al., 1994; Krömer,1995; Gardeström and Lernmark, 1995;Gardeström, 1996; Hoefnagel et al., 1998;Padmasree and Raghavendra, 1998, 2000;Atkin et al., 2000b; Gardeström et al., 2002).

Most of the work on the interaction ofmitochondrial respiration and chloroplastphotosynthesis is based on the use of meta-bolic inhibitors that inhibit specific reac-tions at low concentrations. An inherentdisadvantage, however, is that these inhibi-tors cause nonspecific effects particularly athigher concentrations (see Section VI).Among the mitochondrial inhibitors referredfrequently in this review are rotenone (an

inhibitor of complex I in the mitochondrialrespiratory chain); antimycin A (an inhibi-tor of complex III); KCN/NaN3 (inhibitorsof complex IV); oligomycin (an inhibitor ofcomplex V); salicylhydroxamic acid(SHAM)/propyl gallate (inhibitors of alter-native oxidase); and aminoacetonitrile(AAN, an inhibitor of glycine decarboxy-lase).

B. Differential Effects on CO 2

Efflux/O 2 Uptake

The occurrence of mitochondrial respi-ration in light has been a matter of debate, fora long time, because of ambiguous reports onthe extent and pattern of dark respiration inlight (Graham, 1980; Raghavendra et al.,1994; Krömer, 1995; Villar et al., 1995; Atkinet al., 1997; Hoefnagel et al., 1998; Atkin etal., 2000a). Some studies have indicated thatdark respiration was either unaffected orstimulated, while others found that respira-tion was inhibited (Table 1). Such a largevariation in these reports appears to be due toa combination of factors: the component ofdark respiration being monitored (CO2 efflux/O2 uptake), the experimental technique be-ing used, and the subject of experimentalsystem (leaves, algal cells, or cell cultures).

It is difficult to monitor precisely CO2/O2 exchange in light by conventional meth-ods because the measurements are compro-mised by the occurrence of related phenom-ena besides dark respiration, for example,photorespiration, photosynthesis, Mehlerreaction, chlororespiration. Each one of theabove processes contributes significantly tothe net CO2 or O2 exchange.

A promising solution was provided bythe technique of mass spectrometry, whichcould distinguish between the uptake/effluxof O2 or CO2 occurring simultaneously dur-ing respiration, photosynthesis, and related

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cellular processes. Mass spectrometric stud-ies using 13/12CO2 and 18/16O2 have revealedthat light has a differential effect on CO2

efflux and O2 uptake (Avelange et al., 1991;Raghavendra et al., 1994; Xue et al., 1996;Atkin et al., 2000a). On illumination, CO2

efflux is suppressed by almost 81%, whileO2 consumption is either unaffected or stimu-lated up to 3.5-fold (Table 2). A major ob-servation from these mass spectrometric ex-periments is that mitochondrial oxidativeelectron transport continues to be active,

irrespective of illumination. The sustenanceof active mitochondrial oxidative electrontransport is essential for optimal photosyn-thesis.

C. Light-Enhanced DarkRespiration (LEDR)

Although the extent of respiration in lightis often debated, the stimulation of dark res-

TABLE 1The Effect of Light on Dark Respiration in Plant Tissues, Determined by DifferentTechniques, Indicating a Large Variation in the Extent of Inhibition

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piration due to illumination, particularly ingreen tissues, is now well established. Therespiratory O2 uptake in dark increases quitesignificantly, soon after illumination (Figure 1).This phenomenon termed as ‘light enhanceddark respiration’ (LEDR) occurs after evenshort periods of exposure to light (Padmasreeand Raghavendra, 1998). The phenomenonof LEDR has been recorded in different ex-perimental systems and the extent of stimu-lation by light, varied from 1.2- to 7-fold(Table 3).

The extent of LEDR is positively corre-lated with the intensity and duration of thepreceeding period of illumination (Raghavendraet al., 1994; Xue et al., 1996). The sensitivity ofLEDR to DCMU (an inhibitor of photosystemII electron transport) and D,L-glyceraldehyde

(inhibitor of Calvin cycle) establishes that LEDRis dependent on products of photosyntheticcarbon assimilation and electron transport(Reddy et al., 1991). Exposure of Euglena gra-cilis, a flagellate to UV radiation decreasedboth the rate of photosynthesis and LEDR,especially at higher light intensities (Ekelund,2000).

LEDR is different from photorespiratorypost-illumination burst (PIB). The phenom-enon of PIB results due to CO2 released dur-ing decarboxylation of photorespiratory gly-cine. In tobacco leaf PIB occurs within 20 safter the light is switched off, while LEDRoccurs between 180 to 250 s after stoppingillumination (Atkin et al., 1998). Further at2% O2 (where photorespiration is minimized)PIB is not seen, whereas LEDR is still ob-

TABLE 2Selected Examples Showing the Differential Effect of Light on Respiratory CO 2 Releaseand O2 Uptake, as Determined by Mass Spectrometry. On Illumination, the DecarboxylationReactions Are Usually Inhibited Resulting in a Decrease of CO 2 Release, While the Processof Oxidative Electron Transport (Indicated by O 2 Uptake) Is Either Unaffected or EvenEnhanced

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served (Atkin et al., 1998; Atkin et al., 2000a).LEDR is also insensitive to AAN (an inhibi-tor of mitochondrial glycine metabolism)demonstrating that LEDR is not directly re-lated to photorespiration (Gardeström et al.,1992; Raghavendra et al., 1994).

The occurrence of LEDR within a fewminutes suggests that the interaction be-tween photosynthesis and respiration is quiterapid and involves primary photosyntheticproducts, especially malate (Raghavendraet al., 1994; Padmasree and Raghavendra,1998). The malate concentration is high atthe end of illumination, and it is rapidlymetabolized during the subsequent darkperiod (Hill and Bryce, 1992). In contrast,

the levels of sucrose, glucose, and fructosedid not change significantly during LEDRin the mesophyll protoplasts of barley (Hilland Bryce, 1992).

Plant mitochondrial pyruvate dehydro-genase complex (PDC) and NAD-malicenzyme are reversibly inhibited in light(see Section I.D). On switching over todarkness, these two enzymes are reacti-vated. Photosynthetically generated malateis oxidized via both malate dehydroge-nase (MDH) and NAD-malic enzyme,resulting in the formation of oxaloacetate(OAA) or pyruvate and CO2. Pyruvate isdecarboxylated by PDC and converted intoacetyl CoA. Both OAA and acetyl CoA

FIGURE 1. LEDR in mesophyll protoplasts of pea. The rate of respirationwas stimulated by three-fold after 15 min of illumination, but not whenprotoplasts were kept in darkness for similar periods. (Modified fromReddy et al., 1991.)

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enter TCA cycle and enhance the processof respiratory CO2 release and oxidativeelectron transport. All these result inan upsurge in CO2 release or O2 uptaketermed LEDR (Raghavendra et al., 1994;Padmasree and Raghavendra, 1998; Atkinet al., 2000a). Thus, malate could be thesubstrate and signal for LEDR, whichcould accomplish the rapid conversion ofphotosynthetically generated reducingpower into ATP. Experiments designed toalter chloroplast malate production or theimport of malate into mitochondria, forinstance, by altering the function of chlo-roplastic MDH or the overexpression/supression of mitochondrial OAA/malatetranslocator would help throw more lighton the cause of LEDR.

D. Mitochondrial Respiration inLight: Modified TCA Cycle

Dark respiration consists of three steps:(1) glycolysis in the cytosol, (2) the TCAcycle, consisting of decarboxylation of car-bon compounds resulting in the productionof NADH/FADH and CO2, (3) the electrontransport chain involving NADH/FADHoxidation to produce ATP, and O2 consump-tion.

The processes of CO2 production andO2 uptake are not as tightly coupled in lightas in darkness. As discussed in the previoussection, there is usually a reduction in theextent of respiratory CO2 efflux, while oxy-gen uptake is either unaffected or even stimu-lated (Table 2). The main reason for such

TABLE 3The Occurrence and Extent of Light Enhanced Dark Respiration (LEDR) in Plant Tissues.The Rate of Respiration Increases Markedly After a Few Minutes of PhotosyntheticCarbon Assimilation and LEDR Is Represented as the % Increase in Respiration JustAfter Illumination Over the Rate of Respiration (Steady State) in Continuous Darkness

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decrease in CO2 release is the partial inhibi-tion/modification of TCA cycle activityoccurring in light. Two of the possible causesthat downregulate/modify TCA cycle activ-ity are (1) reversible inactivation of mito-chondrial pyruvate dehydrogenase complexin light (Budde and Randall, 1990), (2) rapidexport of oxoglutarate out of mitochondria(Hanning and Heldt, 1993), thus resultingin a short-circuit of Kreb’s cycle.

PDC is phosphorylated (inactivated) inlight by a PDC-protein kinase and dephos-phorylated (activated) in darkness (Leuthyet al., 1996; Randall et al. 1996). The inac-tivation of PDC is linked to photosyntheticactivity as indicated by its sensitivity toDCMU (inhibitor of PSII) and the absenceof the phenomenon in etiolated seedlings.Further, the products of glycine decarboxy-lation, NADH and NH4

+, also enhance thephosphorylation of PDC. Conditions thatreduce photorespiration (high CO2 and/orlow O2) limit the extent of PDC inactiva-tion. Taken together these reports indicatethat under conditions of high rates of photo-synthesis or photorespiration, PDC is inac-tivated, but oxidation of glycine or malatecontinues.

The second possible reason for reducedCO2 efflux in light is the export of TCAcycle compounds from mitochondria tochloroplast (mainly for NH4+ assimilationin light), thus limiting the substrates avail-able for further steps of the TCA cycle.Mitochondria export TCA cycle interme-diates in the form of citrate, which is con-verted into oxoglutarate in cytosol and sentinto chloroplast (Figure 2; Chen and Gadal,1990; Gout et al., 1993; Hanning and Heldt,1993; Krömer, 1995; Atkin et al., 2000b).This results in a partial activity of TCAcycle in light resulting in reduced CO2

efflux (Parnik and Keerberg, 1995). Thusthe main function of TCA cycle in lightseems to be the supply of carbon skeletonsto the chloroplasts for ammonium assimi-

lation (Weger et al., 1988 and Weger andTurpin, 1989).

The ATP levels in cytosol during lightdo not appear to play any crucial role indownregulating TCA cycle. The ATP/ADPratio required in the cytosol to decreasemitochondrial respiration is much higherthan that usually occurs. In fact, the ATP/ADP ratio in cytosol is lower in light com-pared to darkness at saturating CO2 levels(Krömer, 1995; Atkin et al., 2000b). If mi-tochondrial function in light were to be in-hibited due to high ATP levels in cytosol, itwould have also inhibited O2 uptake besidesCO2 release. However, experimental evi-dence shows that O2 uptake does not de-crease much in light. The mitochondrialelectron transport chain oxidizes not onlythe NADH produced by the partially activeTCA cycle, but also that produced byphotorespiratory glycine decarboxylation.Glycine oxidation therefore can contributesignificantly to oxygen consumption bymitochondria in light.

II. ESSENTIALITY OFMITOCHONDRIAL RESPIRATIONFOR PHOTOSYNTHESIS

One of the first indications about theimportance of mitochondria came from thefrequent observation of a positive relation-ship between dark respiration and photosyn-thesis. A strong positive correlation existsbetween steady state rates of photosynthesisand respiration in a wide range of species(Ceulemans and Saugier, 1991) and the res-piratory rate of leaves increases significantlyafter the light period or hours of illumination(Raghavendra et al., 1994; Atkin et al., 1998).Mitochondrial oxidative metabolism (particu-larly oxidative electron transport and oxida-tive phosphorylation) has been shown to beessential for maintaining high rates of photo-

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synthesis under a variety of conditions. Thebeneficial effects of mitochondria are knownduring not only short cycles of darkness andillumination, but also under stress environ-ments like photoinhibitory light or low tem-perature (Vani et al., 1990; Saradadevi andRaghavendra 1992; Saradadevi et al., 1992;Shyam et al., 1993; Hurry et al., 1995).

The marked interaction between photo-synthesis and respiration is convincinglydemonstrated by experiments with leaf me-sophyll protoplasts and intact leaves (Krömeret al., 1988; Vani et al., 1990; Krömer andHedlt, 1991a; Krömer et al., 1993; Padmasreeand Raghavendra, 1999a,b,c) using inhibi-tors of mitochondrial metabolism. The es-sentiality of different components of mito-chondrial respiration for chloroplasticphotosynthesis is documented by the use oftypical mitochondrial inhibitors. For example,oligomycin was employed as inhibitor ofoxidative phosphorylation (Krömer et al.,1988; Krömer and Heldt, 1991a; Krömer etal., 1993), antimycin A to inhibit the cyto-chrome pathway (Igamberdiev et al., 1997a,b;Padmasree and Raghavendra, 1999a,b,c),while SHAM was used to inhibit the alterna-tive pathway (Igamberdiev et al., 1997a,b;Padmasree and Raghavendra, 1999a,b,c). Onthe other hand, the importance of glycolyticreactions and TCA cycle in stimulating pho-tosynthetic O2 evolution was assessed by theusage of sodium fluoride and sodium mal-onate, respectively (Vani et al., 1990). Re-cently, studies were carried out using mu-tants deficient in mitochondrial glycinedecarboxylase (Igamberdiev et al., 2001).

A. Restriction of CO 2

Assimilation, but Not ofPhotochemical Activities

Preliminary experiments have indicatedthat the inhibitors employed during these

studies had no direct effect on chloroplasts(Krömer et al., 1988; Padmasree andRaghavendra, 1999a). The suppression ofphotosynthetic activity was reversed and thefull rate was restored when the protoplastswere ruptured, leaving the chloroplasts in-tact. These results indicate that the stronginhibition of photosynthesis observed witholigomycin or antimycin A or SHAM wasdue to not an effect on chloroplast photo-synthesis as such, but interference of reac-tions between the chloroplasts, cytosol andmitochondria (Figure 3).

The statistical significance of the inter-action between photosynthesis and respira-tion only in the presence of CO2 (but not inits absence) suggested that carbon assimila-tion was a prerequisite (Vani et al., 1990). Itis intriguing to note that despite the smalleffects of SHAM on respiration or ATPlevels, the decrease in photosynthetic activ-ity is always pronounced (Padmasree andRaghavendra, 1999a).

A recent comprehensive study reexam-ined the effects of mitochondrial inhibitors:oligomycin, antimycin A, and SHAM onthe photosynthetic carbon assimilation andphotochemical electron transport activities,monitored in intact mesophyll protoplasts(Padmasree and Raghavendra, 2001b).When mesophyll protoplasts were illumi-nated in presence of mitochondrial inhibi-tors, there was a significant decrease (>45%)in HCO3

–-dependent O2 evolution, while thedecrease in O2 evolution was marginal(<10%) in presence of benzoquinone (BQ),[PSII mediated] and NO2– [dependent onPSII + PSI] as electron acceptors (Figure 4).DCMU, a typical photosynthetic inhibitordecreased drastically all the three reactions:HCO3

– or BQ or NO2–-dependent O2 evolu-

tion in mesophyll protoplasts. The effect ofmitochondrial inhibitors on photosyntheticreactions was similar in the presence orabsence of NH4Cl, an uncoupler, indicatingthat photophosphorylation also was not af-

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fected. Thus, mitochondrial oxidative me-tabolism (through both cytochrome and al-ternative pathways) was essential for themaintenance of photosynthetic carbon as-similation, but had no direct effect on PSI-or PSII-dependent photochemical electrontransport activities in mesophyll protoplastsof pea. However, during a long-term incu-bation, interference with mitochondrial me-tabolism can lead to disturbance in photo-chemical activities through feedback effectsof thylakoid overenergization (Krömer etal., 1993).

B. Importance at Both Limitingand Optimal CO 2

The rate of photosynthetic O2 evolutiondepends on not only light intensity but alsothe CO2 concentration. At saturating CO2

and light, the rate of photosynthesis is lim-ited by the flux of assimilated carbon intosucrose and at limiting CO2 and saturatinglight, the rate of photosynthesis is limited

by rubisco activity (Krömer et al., 1993). Atoptimal CO2 (nonphotorespiratory condi-tions), the photosynthetic demand for ATPis expected to be very high, while such aneed for ATP would be lower at limitingCO2. The decrease in the rate of photosyn-thesis due to mitochondrial inhibitors, oli-gomycin, antimycin A, or SHAM at opti-mal CO2 (1.0 mM NaHCO3) was muchstronger than that at limiting CO2 (0.1 mMNaHCO3) under similar conditions (Figure 5).Nevertheless, the significant decrease in therate of photosynthesis under both limitingand optimal CO2 in the presence of theseinhibitors suggests that mitochondrial oxi-dative metabolism is essential for maximalphotosynthesis at both limiting CO2

(photorespiratory conditions) as well asoptimal CO2 (Padmasree and Raghavendra,1999a).

At optimal CO2, most of the photosyn-thate is converted to sucrose-consuming ATPand Glc-6-P. Both oligomycin and antimycinA while causing a decrease in photosynthesisalso raised the levels of Glc-6-P and triose-Pat optimal CO2 (Krömer et al., 1988; Krömer

FIGURE 3. Marked suppression of bicarbonate dependent O2 evolution (�) in pea mesophyllprotoplasts by both antimycin A (inhibitor of cytochrome pathway of mitochondrial electron trans-port) and SHAM (inhibitor of alternative pathway). These two compounds had no direct effect onchloroplast photosynthesis (∆). (Adapted from Padmasree and Raghavendra, 1999a.)

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FIG

UR

E 4

. C

hang

e in

rat

es o

f ox

ygen

evo

lutio

n w

hen

mes

ophy

ll pr

otop

last

s ar

e in

cuba

ted

with

diff

eren

tco

ncen

trat

ions

of

olig

omyc

in,

antim

ycin

A,

DC

MU

or

SH

AM

. �

: H

CO

3–-d

epen

dent

O2

evol

utio

n (r

efle

cts

the

tren

d of

CO

2 a

ssim

ilatio

n);

�:

Rat

es o

f ox

ygen

evo

lutio

n w

hen

elec

tron

s ar

e tr

ansf

erre

d fr

om H

2O

to

NO

2

(rep

rese

ntin

g P

SII

and

PS

I ac

tivity

); ∆

: B

enzo

quin

one-

depe

nden

t ox

ygen

evo

lutio

n (in

dica

tes

PS

II ac

tivity

excl

udin

g P

SI,

H2O

, to

BQ

). In

con

trol

s, th

e ra

tes

of O

2 ev

olut

ion

in µ

mol

mg–1

Chl

h–1

und

er d

iffer

ent c

ondi

tions

wer

e 15

4 (H

CO

3– -

depe

nden

t O

2 ev

olut

ion)

; 44

5 (B

Q-d

epen

dent

O2

evol

utio

n) o

r 52

(N

O2-d

epen

dent

O2

evol

utio

n).

(Ada

pted

fro

m P

adm

asre

e an

d R

agha

vend

ra,

2001

b.)

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FIG

UR

E 5

. E

ffect

of

antim

ycin

A (

A)

or S

HA

M (

B)

on p

hoto

synt

hesi

s at

opt

imal

(1.

0 m

M N

aHC

O3)

or li

miti

ng (

0.1

mM

NaH

CO

3)

CO

2 in

mes

ophy

ll pr

otop

last

s of

pea

. (M

odifi

ed f

rom

Pad

mas

ree

and

Rag

have

ndra

, 19

99a.

)

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and Heldt, 1991a; Krömer et al., 1993;Padmasree and Raghavendra, 1999a). Obvi-ously, mitochondrial metabolism and ATPsupply are essential for the maintenance ofsucrose biosynthesis. At limiting CO2 there iseither no change or only a marginal decreasein Glc-6-P in the presence of oligomycin orantimycin A (Padmasree and Raghavendra,1999a). However, the presence of SHAM(which leads to only a limited ATP produc-tion) caused a decrease in the levels of Glc-6-Pand had no effect on ATP/ADP levels,while markedly affecting photosynthesis inmesophyll protoplasts (Padmasree andRaghavendra, 1999a). Thus, the mitochon-drial supply of ATP for sucrose synthesis maybe only a secondary factor during the interac-tion with photosynthesis at limiting CO2.

Although the marked decrease in the rateof photosynthetic O2 evolution at both opti-mal and limiting CO2 demonstrates the es-sentiality of mitochondrial oxidative metabo-lism in optimizing photosynthesis, underphotorespiratory conditions mitochondrialelectron transport is more crucial than oxida-tive phosphorylation in benefitting photosyn-thesis. A major function of the mitochon-drion in a photosynthesizing cell, particularlyunder low light intensities and optimal CO2,seems to be the supply of ATP for cytosoliccarbon metabolism, that is, sucrose synthe-sis. In high light, mitochondria take on theadditional role of oxidizing the excess reduc-ing equivalents generated by photosynthesis,preventing overreduction of chloroplasticredox carriers and thus maintaining high ratesof photosynthesis (Krömer, 1995; Padmasreeand Raghavendra, 1998, 2000).

C. Role of Cytochrome andAlternative Pathways ofOxidative Electron Transport

The sensitivity of protoplast photosyn-thesis to mitochondrial inhibitors at limiting

CO2 (when the ATP requirement for photo-synthesis is expected to be low) and the lackof correlation between the photosyntheticrates and the ratios of ATP/ADP in proto-plasts (particularly in presence of SHAM)have indicated that mitochondrial electrontransport activity is more important than theoxidative phosphorylation for optimal pho-tosynthesis (Padmasree and Raghavendra,1999a).

Plant mitochondria have the unique ca-pability of oxidizing NADH/FADH throughtheir electron transport by two differentroutes: (1) cyanide-sensitive cytochromepathway and (2) cyanide-resistant alterna-tive pathway (Lambers, 1985; Vanlerbergheand McIntosh, 1997; Mackenzie and McIn-tosh, 1999). The alternative pathway is cata-lyzed by alternative oxidase (AOX), whichhas been purified, characterized, and its geneisolated (Siedow and Umbach, 2000). Whilethe molecular biology and regulation ofAOX are studied in detail (McIntosh, 1994;Siedow and Umbach, 1995, 2000), the in-formation on the physiological significance/metabolic function of AOX is still limited.

The relative proportion of cytochromeand alternative pathways is flexible andvaries with environmental conditions suchas temperature, age of the tissue, and injury/wounding. The circumstantial evidence sug-gests that the operation of alternative path-way is likely to increase in illuminated planttissues. The levels of AOX increase duringgreening of etiolated leaves (Atkin et al.,1993). The accumulation of sugars duringthe illumination promotes the engagementof alternative pathway (Azcón-Bieto, 1992).A significant part of the light-enhanced darkrespiration (LEDR) appears to involve al-ternative pathway (Igamberdiev et al.,1997a). It is not known, however, if there isany modulation by illumination of the ex-tent of mitochondrial electron transportthrough the alternative pathway.

Although the essentiality of mitochon-drial oxidative phosphorylation for photo-

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synthetic carbon assimilation is well estab-lished, the role of cytochrome and alterna-tive pathways in benefitting photosyntheticmetabolism is examined only to a limitedextent. The importance of cytochrome andalternative pathways during photosynthesiswas studied in mesophyll protoplasts of peaand barley, using low concentrationsof mitochondrial inhibitors: oligomycin (in-hibitor of oxidative phosphorylation), anti-mycin A (inhibitor of cytochrome pathway)and salicylhydroxamic acid (SHAM, an in-hibitor of alternative pathway). All thethree compounds decreased the rate of pho-tosynthetic O2 evolution in mesophyll pro-toplasts, but did not affect chloroplast pho-tosynthesis (Krömer et al., 1988; Krömerand Heldt, 1991a; Krömer et al., 1993;Igamberdiev et al., 1997a, 1998; Padmasreeand Raghavendra, 1999a,b,c). The markedsensitivity of photosynthesis to both SHAMand antimycin A suggests that the alterna-tive pathway is as essential as the cyto-chrome pathway for optimal photosynthe-sis. These results also demonstrate animportant role of the alternative pathway inplant cells: essentiality for chloroplast pho-tosynthesis.

The importance of the alternative path-way during the interaction between respira-tion and photosynthesis is suggested by alsothe sensitivity of LEDR to SHAM in meso-phyll protoplasts of barley (Igamberdiev etal., 1997a) and algae Selenastum minutum,Chlamydomonas reinhardtii, and Euglenagracilis (Lynnes and Weger, 1996; Xue etal., 1996; Ekelund, 2000).

Restriction of cytochrome pathway byantimycin A or uncoupling of cytochromepathway from oxidative phosphorylation byoligomycin prolonged both the inductionphase of photosynthesis and the activationof NADP-MDH during transition from darkto light (Igamberdiev et al., 1998; Padmasreeand Raghavendra, 1999b). The exact mecha-nism of the optimization of photosynthesis

by alternative pathway is not completelyunderstood, but one of the reasons appearsto be the effective modulation of intracellu-lar redox state (Padmasree and Raghavendra,1999c). A major function of AOX pathwayin mesophyll cells can be the maintainanceof the oxidation of malate, particularly un-der excess light (see Section IV.B).

A recent report suggests that the phe-nomenon of the Kok effect (progressive lightinduced inhibition of dark respiration at lowlight intensities) is modulated strongly bycytochrome pathway of mitochondrial elec-tron transport. The alternative pathway ap-pears to be less important in modulating theKok effect (Padmavathi and Raghavendra,2001).

D. Pronounced Interaction inAlgal Mutants/Guard Cells

The interaction between respiration andphotosynthesis is quite pronounced in cellsthat are deficient in Rubisco/Calvin cycleactivity, such as stomatal guard cells andmutants of Chlamydomonas (Raghavendraet al., 1994).

Guard cells have high rates of respiratoryactivity but contain very low levels of Rubiscoand consequently limited carbon metabolismthrough Calvin cycle (Raghavendra and Vani,1989; Parvathi and Raghavendra, 1995).Despite the limited CO2 fixation in guardcells, the reduced equivalents produced bytheir chloroplasts are exported to the cytosolthrough OAA-malate or PGA-DHAP shuttles(Shimazaki et al., 1989). The reduced pyri-dine nucleotides formed in the cytosol fromthe oxidation of malate and/or DHAP mayact as the respiratory substrates for mito-chondrial ATP production needed for K+

uptake. A very strong interaction betweenrespiration and photosynthesis has beenshown in guard cell protoplasts of Vicia faba

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and Brassica napus at varying O2 concentra-tions (Mawson, 1993). A strong cooperationbetween chloroplasts and mitochondria ap-pears to be essential for the maintenance ofguard-cell bioenergetic processes.

A similar situation appears to operate intwo mutants of Chlamydomonas reinhardtii,one devoid of Rubisco and the other lackingfunctional chloroplast ATP synthase. TheC. reinhardtii mutant FUD50 lacks theβ-subunit of chloroplast ATP synthase andcannot produce ATP during photophospho-rylation (Gans and Rébéille, 1988). A modi-fied strain of this mutant FUD50su can growunder photoautotrophic conditions, althoughit still showed no synthesis of the β-subunitof the coupling factor. Photosynthesis inFUD50su mutant was extremely sensitiveto inhibitor antimycin A, a specific inhibitorof mitochondrial electron transport. Photo-synthesis in the FUD50su strain is achievedthrough an unusual interaction betweenmitochondria and chloroplasts (Lemaire etal., 1988). The export of reduced com-pounds, made in light, from the chloroplastto the mitochondria elicits ATP formationin the latter, and ATP is subsequently im-ported to the chloroplast.

III. PROTECTION AGAINSTPHOTOINHIBITION

Photoinhibition can be defined as themarked decrease in the photosynthetic rateunder supraoptimal light or limitation onCO2 assimilation. Such situations developwhen there is excess light or conditions lim-iting the biochemical reactions, for example,low temperature, water stress, limiting CO2,limiting N2, or any limitation on enzymes.The function of photochemical electrontransport is optimized when the reductantsgenerated in light are quickly used up forbiochemical reduction of carbon (or nitro-

gen or sulfur). Any imbalance between thephotochemical and biochemical processesleads to the phenomenon of photoinhibition(Long et al., 1994; Andersson and Barber,1996).

Photoinhibition occurs due to theoverreduction of the photosynthetic elec-tron transport system. Reactive oxygen spe-cies generated in the light as a result of theMehler reaction can lead to damage of thephotochemical apparatus, particularly PS II.The chloroplasts have the necessary ma-chinery to repair the damage caused to PSII(Carpentier, 1997; Critchley, 1998; Ohad etal., 2000). The recovery is accomplished bya continuous synthesis of PSII components,particularly D1 protein. However, such re-covery is optimal under very low light andis rather slow at moderate light intensities(Park et al., 1996; Singh et al., 1996; Ander-son, 2001). Thus, photoinhibition of photo-synthesis sets in when the rate of damageexceeds that of repair (Long et al., 1994;Critchley, 1998).

Plants have evolved in different ways tocope with photoinhibition by preventive aswell as repair mechanisms. Some of themare adjustment of chloroplast antennae size,xanthophyll cycle, CO2 fixation, photores-piration, water-water cycle, PS I cyclic elec-tron transport, scavenging reactive moleculesthrough antioxidant enzymes, rapid turn-over of D1 protein of PS II (Niyogi, 1999).However, in the past decade there has beenconvincing evidence to show that mitochon-drial respiration, especially oxidative elec-tron transport and phosphorylation, playa significant role in protecting the chloro-plasts from photoinhibition (Saradadevi andRaghavendra, 1992; Shyam et al., 1993;Raghavendra et al., 1994; Singh et al.,1996; Padmasree and Raghavendra, 1998;Atkin et al., 2000b). The protection of chlo-roplast photosynthetic machinery againstphotoinhibition is accomplished by mito-chondria through not only the oxidative elec-

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tron transport/oxidative phosphorylation butalso through the key photorespiratory reac-tions.

A. Oxidative Electron Transportand Oxidative Phosphorylation

Even at very low concentrations, anti-mycin A or sodium azide or oligomycin en-hanced markedly the extent of photoinhibitionin mesophyll protoplasts of pea (Saradadeviand Raghavendra, 1992). These inhibitiors atsuch low concentrations did not affect photo-synthesis directly. Sodium fluoride (inhibitorof glycolysis) or sodium malonate (inhibitorof TCA cycle) did not significantly affectphotoinhibition (Table 4). Apparently, oxi-dative electron transport and phosphoryla-

tion play a major role in protecting photosyn-thesis aganist photoinhibition.

After an initial increase, dark respirationdecreases significantly after prolonged expo-sure to photoinhibitory light in pea mesophyllprotoplasts (Saradadevi and Raghavendra,1992) or algal cells of Anacystis nidulans andChlamydomonas reinhardtii (Shyam et al.,1993; Singh et al., 1996). These observationsindicate a marked correlation between chloro-plast and mitochondrial activity during evenphotoinhibition. The initial increase mightrepresent an enhanced oxidation of excessredox equivalents generated by chloroplastunder high light. A subsequent decrease isprobably due to the reduced flux of redoxequivalents from chloroplasts, which are nowphotoinhibited.

The initial increase in respiration oc-curred even in the presence of KCN in

TABLE 4A Comparison of the Effect of Five Metabolic Inhibitors on Respiration, Photosynthesis,and Photoinhibition in Mesophyll Protoplasts of Pea. The Protoplasts Were Examined forTheir Photosynthetic Activity After a Preincubation (With or Without Inhibitors) for 10min at 30 oC in Either Darkness or Photoinhibitory Light (Adapted from Saradadevi andRaghavendra, 1992)

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Chlamydomonas reinhardtii (Singh et al.,1996), implying that it mostly representsthe activity of the alternative pathway (Singhet al., 1996). The alternative pathway ofmitochondrial oxidative electron transportis a potential channel for “overflow” anddissipation of excess photosynthetic reduc-tants (Lambers, 1982; Millar and Day, 1997).Alternative pathway therefore is likely toplay a significant part in preventing theoverreduction of chloroplast photosyntheticapparatus and to alleviate photoinhibition.Further experiments are needed to confirmthe role of the alternative pathway of mito-chondrial oxidative electron transport inprotection aganist photoinhibition.

B. Sustenance of RepairMechanism by MitochondrialATP

Photoinhibition is often a result ofimbalance between the synthesis and deg-radation of D1 protein. Supraoptimal lightaccelerates the degradation while slow-ing down the process of synthesis of D1protein and subsequent recovery (Figure 6).In the cyanobacterium Anacystis nidulansand green alga Chlamydomonas rein-hardtii, inhibition of dark respiration byNaN3 or KCN not only increased photo-inhibition but also accelerated photoin-hibition (Shyam et al., 1993; Singh et al.,1996). The uncoupler FCCP also had asimilar effect of intensifiyng and hasten-ing photoinhibition in both the organ-isms (Shyam et al., 1993; Singh et al.,1996).

In algal cells, mitochondrial respirationmay help in even the recovery of photosyn-thesis after photoinhibition. Treatment withsodium azide or FCCP slowed down recov-ery in Anacystis nidulans (Shyam et al.,1993). Similarly, the use of KCN and FCCP

impaired the reactivation of photosynthesisin Chlamydomonas reinhardtii (Singh et al.,1996). The above results imply that the pro-cess of recovery that involves synthesis ofD1 protein is helped by mitochondrial oxi-dative phosphorylation (Singh et al., 1996).

C. Photorespiratory Reactions

In C3 plants, photorespiration helps inreducing/preventing the damage caused bysupraoptimal light. A classic and convinc-ing demonstration of such a role is providedby transgenic tobacco plants with alteredGS activity. The transgenics with increasedGS2 (a key photorespiratory enzyme) activ-ity had higher photorespiratory rates andmore tolerance to supraoptimal light thanthe wild-type plants. On the other hand,those with reduced GS2 were low on photo-respiration and were sensitive to high light(Kozaki and Takeba, 1996).

There are two possible ways by whichphotorespiration could help preventphotoinhibition under excess light and lim-ited CO2 assimilation: (1) by using up thereducing power generated by photochemi-cal reactions in chloroplasts and (2) main-taining the optimal Pi levels in chloroplasts.When stomata are closed (e.g., drought) orwhen CO2 is limiting, disturbance in thelevels of NADPH and ATP is prevented byregulatory mechanisms, which include pho-tosynthetic control and photorespiratoryglycolate metabolism (Osmond et al.,1997). Photorespiration was shown to beessential and even more important than theMehler or Asada reactions in preventingphotoinactivation of photosynthesis in Che-nopodium bonus-henricus (Heber et al.,1996).

When protoplasts of barley mutants withreduced glycine decarboxylation were incu-bated in limiting CO2, their chloroplasts had

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FIG

UR

E 6

. Im

port

ance

of m

itoch

ondr

ial o

xida

tive

elec

tron

tran

spor

t for

pho

tosy

nthe

sis

durin

g ph

otoi

nhib

ition

or r

ecov

ery

afte

r ph

otoi

nhib

ition

in A

nacy

stis

nid

ulan

s. (

A)

Effe

ct o

f pho

toin

hibi

tory

ligh

t (25

00 µ

mol

m–2

s–1

) on

pho

tosy

nthe

sis

in th

eab

senc

e (∆

) or

pre

senc

e (∆

) of

1 m

M s

odiu

m a

zide

. (B

) R

eact

ivat

ion

of p

hoto

synt

hesi

s fr

om p

hoto

inhi

bitio

n in

the

abse

nce

or p

rese

nce

of 1

mM

sod

ium

azi

de.

50%

pho

toin

hibi

ted

cultu

res

wer

e us

ed t

o st

udy

the

reac

tivat

ion

ofph

otos

ynth

esis

fro

m u

nder

dim

ligh

t. (M

odifi

ed f

rom

Shy

am e

t al

., 19

93.)

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high ratios of ATP/ADP and NADPH/NADP (Igamberdiev et al., 2001). This in-dicates that glycine decarboxylation andassociated NH+4 recycling are sinks for ex-cess chloroplastic reductants and help toprevent a overreduction of the chloroplast.The GDC-deficient barley mutants alsoshowed significant increase in the activityof malate valve and chloroplastic NADP-MDH, apart from an increase in the activityof NAD-MDH of cytosol and mitochon-dria. Obviously, the malate shuttle compen-sates for the decreased glycine decarboxy-lation by dissipating the excess reducingequivalents (Gardeström et al., 2001).

In cotton leaves, maintained at low O2

concentration, a nonphotorespiratory con-dition, photosynthesis, was severely inhib-ited under strong light, compared with theones kept at normal O2 levels. The low O2samples also had reduced levels of chloro-plastic Pi. When Pi was fed to these leaves,the rates of photosynthesis were restored tothe levels of those kept in normal air con-taining 21% O2. This indicates that Pi limi-tation could partly be alleviated byphotorespiratory recycling of Pi. Thus, pho-torespiration reduces photoinhibition bykeeping up rates of photosynthesis throughmaking Pi available for the process (Guo etal., 1995).

Mitochondria, being major players inthe photorespiratory pathway, have to inter-act with chloroplasts and peroxisomes andtake part in balancing the photosyntheticredox equivalents and protection againstphotoinhibition.

D. Significance UnderTemperature or Water Stress

In addition to protection from photo-inhibition, mitochondria also help to opti-mize photosynthesis under stress conditions

like chilling temperature or osmotic stress(Table 5). After a period of cold hardening,the leaves of winter rye exhibited an in-crease in the rates of dark respiration inlight along with those of photosynthesis(Hurry et al., 1995). Oligomycin treatmentresulted in the inhibition of photosynthesismore in cold hardened leaves than that innonhardened ones, suggesting that the in-crease in photosynthetic capacity followingcold hardening is contributed to by mito-chondria. A similar situation of increasedtolerance to photoinhibition following coldhardening has been reported in the leaves ofwinter and spring wheat (Hurry and Huner,1992).

Circumstantial evidence points out tothe possible roles of AOX during the main-tenance of photosynthesis in low tempera-ture. The level of alternative oxidase pro-tein in tobacco (Vanlerberghe and McIntosh,1992) as well as the capacity of alternativerespiration (Rychter et al., 1988) usuallyincrease at low temperatures. The extent ofelectron partitioning to the alternative oxi-dase raises significantly at low tempera-tures in cold grown mung bean (Gonzàlez-Meler et al., 1999). These results indicate arole for alternative respiratory pathway inprotecting the plant tissues from chillingand related photoinhibition and suggest ageneral increase in alternative respirationunder stress conditions.

However, Ribas-Carbo et al. (2000) havefound increased electron flow in the alter-native pathway following cold treatment ina chilling sensitive cultivar of maize com-pared with a chilling tolerant one, indicat-ing no specific role for the alternative path-way of respiration in conferring chillingtolerance. Similarly, no specific increase inalternative respiration ocurred followingchilling in soybean cotyledons (Gonzàlez-Meler et al., 1999). Further studies and di-rect evidence are needed to assign any di-rect role of alternative pathway in the

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protection of photosynthesis from chillingstress in plants.

Mitochondrial respiration seems to berelated to decreased photosynthesis and in-creased susceptability to photoinhibitionunder osmotic stress. Mesophyll protoplastsof pea kept in hyperosmotic medium werehighly susceptable to photoinhibition whenthey were exposed to photoinhibitory light.On exposure to hyperosmotic medium at0°C, both photosynthetic and respiratoryrates decreased, indicating a correlationbetween the two processes (Saradadevi andRaghavendra, 1994). However, at 25°C, res-piration increased, while photosynthesis de-

creased. More experiments are needed tounderstand the role of respiration vis-a-visphotosynthesis during osmotic stress undervarying temperatures.

IV. OPTIMIZATION OFPHOTOSYNTHETIC CARBONASSIMILATION

The optimization of photosynthetic car-bon assimilation requires a coordination ofdifferent components: generation and use ofassimilatory power (ATP and NADPH),

TABLE 5Correlation Between the Pattern of Changes in Respiration and the Extent ofPhotoinhibition in Different Plant Tissues in Presence of Mitochondrial Inhibitors orStress Conditions. Any Decrease in Respiratory Activity Leads to an Increase in theExtent of Photoinhibition, While an Increase in Respiration Helps to Decrease thePhotoinhibition

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induction of photosynthesis, activation ofenzymes, and maintenance of metabolitelevels. In a photosynthesizing cell the mito-chondrial respiratory system may benefitdifferent components of chloroplast photo-synthesis by modulating any of the abovecomponents. However, emphasis has alreadybeen made on the significant role of mito-chondria in maintaining either cytosolic re-dox status or ATP or both (Krömer et al.,1988; Krömer and Heldt, 1991a; Krömer etal., 1993; Raghavendra et al., 1994;Gardeström and Lernmark, 1995; Krömer,1995; Igamberdiev et al., 1998; Padmasreeand Raghavendra, 1998).

A. Sustenance of SucroseBiosynthesis: Role of ATP

The biosynthesis of sucrose, one of themajor end products of photosynthetic car-bon assimilation, occurs in the cytosol ofmesophyll cells. Sucrose biosynthesis in thecytosol requires a continuous supply of car-bon skeletons and energy. Although it isobvious that chloroplasts play a significantrole in supplying the carbon compounds forthe synthesis of sucrose, the relative impor-tance of mitochondria in meeting the cyto-solic demands for ATP, particularly duringsucrose formation is emphasized only dur-ing the last decade (Raghavendra et al., 1994;Gardeström and Lernmark, 1995; Krömer,1995; Hoefnagel et al., 1998; Padmasreeand Raghavendra, 1998; Atkin et al., 2000b).Studies with a starchless mutant NS 458 ofNicotiana tabacum (defective in plastidphosphoglucomutase) in the presence of oli-gomycin also suggested that the mitochon-drial supply of ATP could affect assimilatepartitioning into sucrose and thereby modu-late photosynthesis (Hanson, 1992).

The transfer of redox equivalents gener-ated during the oxidation of TCA cycle in-

termediates along the mitochondrial elec-tron transport chain accumulate significantamounts of ATP in the mitochondrial ma-trix. Mitochondria have a very high capac-ity for ATP synthesis, in fact higher thanthat of chloroplasts, producing up to 3 ATPper NAD(P)H compared with 1.5 to 2.0ATP per NAD(P)H in the chloroplast(Hoefnagel et al., 1998; Siedow and Day,2000). It is possible that the ATP poolsgenerated in the mitochondrial matrix aretranslocated to cytosol (through adenylatetranslocator) to be used in sucrose synthesisor even imported into chloroplasts to beused in various other biosynthetic processeslike protein synthesis, NH4+ assimilation,metabolite transport, and maintenance(Hoefnagel et al., 1998).

It is possible in light that mitochondrialrespiration is subjected to adenylate control(Hoefnagel et al., 1998). However, the abil-ity of plant mitochondria to switch betweenthe rotenone-sensitive and rotenone-insen-sitive as well as the cyanide-sensitive cyto-chrome pathway and cyanide-resistant al-ternative pathways provides for a flexiblesystem and ATP production. However, thedegree to which mitochondrial ATP supplyin the light required for optimal photosyn-thesis depends on the balance of ATP pro-duction and consumption in chloroplasts.Two key observations indicate the primaryrole of mitochondria in assisting chloro-plasts in meeting the cytosolic demands ofATP for sucrose synthesis: (1) An increasein cytosolic or cellular levels of glucose-6-P and other phosphates (e.g., fructose-6-Pand fructose-2,6-bisphosphate) in the pres-ence of oligomycin or antimycin A (Krömerand Heldt, 1991a; Krömer et al., 1993,Padmasree and Raghavendra, 1999c). Sub-cellular analysis of protoplasts revealed thatthe increase in hexose monophosphates wasmostly in the cytosol, demonstrating therestriction of sucrose biosynthesis (Krömeret al., 1992); (2) Restriction of mitochon-

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drial ATP synthesis by oligomycin or an-timycin A or photorespiratory glycine oxida-tion using AAN in isolated protoplasts causeda marked reduction in ATP/ADP ratios in thecytosolic and mitochondrial compartmentsthan that of chloroplasts (Gardeström et al.,1981; Gardeström and Wigge, 1988; Krömerand Heldt, 1991a; Krömer et al., 1993;Igamberdiev et al., 1998).

The change in the levels of intracellularATP and ADP during illumination causedby mitochondrial inhibitors at limiting CO2was in contrast to that of photosynthesis.Despite the expectation that ATP demandswould be low at limiting CO2, there was asteep positive correlation between the ratesof photosynthesis and ratios of ATP to ADPin protoplasts in the presence of oligomycinor antimycin A but not SHAM (Figure 7).The Glc-6-P level increased by about 19 to30% in the presence of both oligomycin andantimycin A at optimal CO2 conditions. The

marked increase in Glc-6-P in mesophyllprotoplasts in the presence of only oligomy-cin or antimycin A but not SHAM suggeststhat the cytochrome pathway of electrontransport (and oxidative phosphorylation)modulates sucrose biosynthesis, while thealternative pathway may not have a signifi-cant role (Padmasree and Raghavendra,1999a).

The restriction of mitochondrial ATPsynthesis by oligomycin and antimycin Awould not only limit sucrose synthesis butalso cause feedback inhibition of photosyn-thetic activity because the phosphatetranslocator in the inner chloroplast mem-brane is regulated by the equilibrium of thetriose-P concentration in the stroma and thecytosol. However, an elevated cytosolic levelof DHAP or reduced flux of DHAP fromchloroplast can also lead to decreased stro-mal PGA level and thereby decreased Calvincycle activity (Krömer et al., 1993,

FIGURE 7. Positive correlation occurs between the ratios of ATP/ADP and the relative rates ofphotosynthesis in pea mesophyll protoplasts in presence of antimycin A (inhibitor of cytochromepathway in mitochondria) but not SHAM, (inhibitor of alternative pathway). In other words, SHAMwhich markedly inhibits photosynthesis of protoplasts, does not alter the relative ratio of ATP/ADP.(Adapted from Padmasree and Raghavendra, 1999a.)

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Padmasree and Raghavendra, 1999a; Flüggeand Heldt, 1991). Thus, mitochondrial oxi-dative electron transport plays a significantrole in optimizing photosynthetic carbonassimilation by sustaining sucrose biosyn-thesis. The flexibility of mitochondrial elec-tron transport chain to meet cytosolic de-mands under both dark and light conditionsmakes it a ready source of energy to meetcellular needs supplementing the chloroplastmetabolism.

B. Maintenance of CellularRedox State: Ratios of Malate/OAA and Triose-P/PGA

Chloroplasts always have a tendency toget overreduced as the rate of photochemi-cal reaction and utilization of reducing po-tential in metabolism have been estimatedto differ by at least 15 orders of magnitude(Huner et al., 1998). It is essential that theexcess reduced equivalents are taken out ordisspiated quickly to prevent damage to thethylakoid membranes (Gillmore, 1997;Niyogi et al., 1998; Niyogi, 1999).

Mitochondria also appear to play a sig-nificant role in maintaining optimal levels ofredox equivalents in the chloroplasts to keepup the Calvin cycle activity, possibly by co-ordinating with peroxisomes and cytosol. Thereductants in excess of the requirements ofthe Calvin cycle are exported out of chloro-plasts through the shuttling of either OAA-malate (by dicarboxylate translocator) orPGA-DHAP (Pi-translocator).

The relative levels of triose-P/PGA andmalate/OAA reflect the redox state of cyto-sol and the cell. Mitochondrial electron trans-port appears to be one of the efficient pro-cesses to use up the reduced equivalents.Any limitation on the mitochondrial me-tabolism leads to a marked rise in the redoxstate of cells, as indicated by the rise in the

ratios of malate/OAA or triose-P/PGA(Padmasree and Raghavendra, 1999c).

The steep gradient in redox levels be-tween stromal compartment and cytosol ismaintained by regulation at several steps suchas (1) chloroplastic NADP-MDH, (2) triose-P/Pi translocator, (3) glycolate/glyceratetranslocator, and (4) glycine/serine translocator(Gardeström et al., 2001). Among these,NADP-MDH functions like a valve, releasingthe excess reductant from chloroplasts asmalate (Scheibe, 1991). Malate valve allowschloroplasts also to provide reducing equiva-lents either to peroxisomes for reduction ofhydroxypyruvate (under photorespiratory con-ditions, Krömer, 1995) or mitochondria to beoxidized by the internal NADH-dehydroge-nase system (under nonphotorespiratory con-ditions; Padmasree and Raghavendra, 1998).This would still allow some of the NADHformed during glycine decarboxylation to beretained in the mitochondria, rather than shut-tling it to the peroxisome to supporthydroxypyruvate reduction. As a result, NADHcan be oxidized within the mitochondria toprovide additional ATP for extrachloroplasticprocesses, such as sucrose synthesis or reduc-tion of PGA in the cytosol (Krömer and Heldt,1991a,b).

The operation of cyanide-resistant alter-native and cyanide-sensitive cytochromepathways of mitochondria appear to be closelyintegrated with the redox regulation duringphotosynthetic metabolism (Padmasree andRaghavendra, 1999c). Restriction of a cya-nide-resistant pathway by SHAM markedlyelevated the malate/OAA ratios, while therestriction of cyanide-sensitive pathway byantimycin A or oligomycin lead to a markedincrease in triose-P/PGA ratios (Figure 8).Because an accumulation of malate repre-sents an overreduction of chloroplasts(Backhausen et al., 1994), the marked in-crease in malate/OAA in the presence ofSHAM suggests an accumulation of redoxpower in protoplasts when AOX pathway is

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FIGURE 8. Changes in the redox state of pea mesophyll protoplasts during photosynthesis in theabsence or presence of typical inhibitors of mitochondrial electron transport. On illumination inpresence of 1.0 mM bicarbonate, the ratios of Triose-P/PGA or Malate/OAA rise with time indicatingthe increase in the redox state of protoplasts. The presence of 250 nM antimycin A (inhibitor ofcytochrome pathway) results in the preferential accumulation of triose-P, while the presence of 200µM SHAM (inhibitor of alternative pathway of mitochondrial electron transport) causes the accumu-lation of malate. (Adapted from Padmasree and Raghavendra, 1999c.)

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restricted. Thus, AOX appears to promotethe consumption of malate in pea mesophyllprotoplasts.

C. Shortening of Induction

The phenomenon of induction (delay inachieving maximal rates) is a common fea-ture of photosynthesis (Edwards and Walker,1983; Walker, 1988). Among the most im-portant factors that cause photosynthetic in-duction are the activation of key chloroplas-tic enzymes (including NADP-malatedehydrogenase, NADP-glyceraldehyde3-phosphate dehydrogenase, stromalFBPase, PRK) and the autocatalytic build-up of Calvin cycle metabolites, for example,

RuBP (Salvucci, 1989; Scheibe, 1991;Edwards and Walker, 1983).

Mitochondrial contribution to photo-synthetic metabolism during photosyn-thetic induction was investigated in me-sophyll protoplasts from barley or pealeaves by using rotenone or oligomycin(Table 6). Both the inhibitors increasedthe lag phase of photosynthetic inductionduring the transition of protoplasts fromdarkness to light (Igamberdiev et al.,1998). Prolongation of photosynthetic in-duction period was observed also withantimycin A, SHAM, and propyl gallate(Figure 9). However, SHAM and propylgallate (inhibitors of alternative pathway)had a negligible effect on the photosyn-thetic induction period (Padmasree andRaghavendra, 1999b).

TABLE 6Prolongation of Photosynthetic Induction in Mesophyll Protoplasts as a Consequence ofRestriction of Mitochondrial Metabolism. The Lag Period for Reaching the Maximum Rateof Photosynthetic Carbon Assimilation (in Presence of 1.0 m M Bicarbonate) is Extendedby the Presence of Mitochondrial Inhibitors

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FIG

UR

E 9

. Pro

long

atio

n of

pho

tosy

nthe

tic in

duct

ion

in p

rese

nce

of ty

pica

l inh

ibito

rs o

f mito

chon

dria

l tra

nspo

rt. T

he la

gpe

riod

for

reac

hing

the

max

imum

rat

e of

pho

tosy

nthe

sis

afte

r sw

itchi

ng o

n th

e lig

ht (

indi

cate

d by

‘L’)

is u

sual

ly le

ss th

an3

min

, w

hile

thi

s la

g in

crea

ses

to a

lmos

t 5

or 8

min

in p

rese

nce

of o

ligom

ycin

(1

µg m

l–1)

or a

ntim

ycin

A (

1 µM

). T

heex

act r

easo

ns fo

r su

ch m

arke

d in

crea

se in

the

phot

osyn

thet

ic in

duct

ion

perio

d ar

e no

t cle

ar. I

t cou

ld b

e al

so o

ne o

f the

cons

eque

nces

of

depr

essi

on i

n ca

rbon

ass

imila

tion

by m

itoch

ondr

ial

inhi

bito

rs.

(Ada

pted

fro

m P

adm

asre

e an

dR

agha

vend

ra,

1999

b.)

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Despite the apparent surplus of ATP inthe chloroplast, the demands for ATP dur-ing the initial phase of photosynthetic in-duction are met by mitochondrial oxidativephosphorylation. The delay by rotenone andoligomycin in photosynthetic induction ap-pears to be caused by a restriction on thereoxidation of redox equivalents from thechloroplasts (and associated reactions) bythe mitochondrial electron transport chain(Igamberdiev et al., 1998). This hypothesismay indeed be complemented by the negli-gible changes in the RuBP levels associatedwith prolonged induction period in the pres-ence of oligomycin and antimycin A in pealeaves (Padmasree and Raghavendra,1999b). The marked sensitivity of photo-synthetic induction period to rotenone orantimycin A suggests that the redox equiva-lents from chloroplasts are being oxidizedby internal dehydrogenase via complex Iand complex III, but not through the roten-one-insensitive dehydrogenases. One of thebenefits of the mitochondrial oxidative elec-tron transport coupled to oxidativephosphorylation is the maintenance or mini-mization of the induction phase of photo-synthesis. The negligible effect on photosyn-thetic induction by SHAM or propyl gallatecompared with the marked delay by antimy-cin A or oligomycin suggests that electrontransport via alternative oxidase may not beas significant as that of cytochrome pathwayduring photosynthetic induction.

Thus, the restriction of mitochondrialactivity leads to an increase in the photo-synthetic induction period in mesophyllprotoplasts of pea as well as barley(Igamberdiev et al., 1998; Padmasree andRaghavendra, 1999b). However, this cor-relation may be incidental because thereis no direct evidence to suggest that re-stricted mitochondrial metabolism is thecause for the prolongation of photosyn-thetic induction.

D. Activation of Enzymes

When leaves are illuminated, the acti-vation state/activity of not only chloroplas-tic enzymes but of several enzymes locatedin different compartments of the cell isstimulated. In this review, attention is drawnto the enzymes involved in coordinating theinteractions between chloroplasts, mitochon-dria, peroxisomes, and cytosol. These en-zymes located in different compartments ofthe cell are fine tuned and coordinated witheach other by specific metabolites.

The light activation of photosyntheticenzymes located in stroma is regulated byseveral factors: ferredoxin-thioredoxin sys-tem, metabolite levels, pH, ionic status, andoxidation-reduction potential (Scheibe, 1990;Faske et al., 1995). Rotenone and oligomy-cin both delayed the activation of chloroplas-tic NADP-MDH during the transition fromdarkness to light (Igamberdiev et al., 1998).The timing of the delay in activation ofNADP-MDH is very similar to the delay inphotosynthetic O2 production and the delayin build-up of nonphotochemical quenching.Restriction of mitochondrial electron trans-port delays alkalization of the chloroplaststroma, which in turn delays the activation ofNADP-MDH and there by the export (andthus use) of redox equivalents (Igamberdievet al., 1998). A marked decrease in lightactivation of not only NADP-MDH, but alsoFBPase, NADP-GAPDH, and PRK in pres-ence of SHAM during steady state photosyn-thesis indicates that the alternative pathwaymay play a significant role in maintaining theactivation status of chloroplastic enzymes(Padmasree and Raghavendra, 2001a). Thisis possible by regulating the redox equiva-lents through malate-OAA shuttle.

Apart from chloroplastic enzymes, cy-tosolic enzymes can also be regulated bylight. One of such is sucrose phosphate syn-

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thase (SPS), which plays a significant roleis sucrose biosynthesis. SPS is subjected toreversible phosphorylation-dephosphoryla-tion cascade. The dephosphorylated enzymeis more active than the phosphorylated from(Huber and Huber, 1996). At high CO2, thedecrease in the mitochondrial and cytosolicATP/ADP ratios caused by the oligomycintreatment at high and low irradiance canlead to a decrease in SPS activity (Krömeret al., 1993). Under high CO2, this inhibi-tion of sucrose synthesis by oligomycin ap-parently increased cytosolic Glc-6-P levelsand caused feedback inhibition of the Calvincycle and photosynthetic activity.

E. Integration withPhotorespiration and NitrogenMetabolism

The interaction of mitochondrial respi-ration, nitrogen metabolism, and photosyn-thesis has been the subject of two extensivereviews (Padmasree and Raghavendra, 1998;Gardestrom et al., 2001). The interplay ofthese three metabolic pathways involvesrecycling of carbon, nitrogen, and markedamounts of reduced equivalents and someATP. The rapidity in the turnover (produc-tion and consumption) of NAD[P]H, reducedferredoxin and ATP allows them to coordi-nate at least four different metabolic path-ways viz., photosynthesis, respiration, pho-torespiration, and nitrogen metabolism.Apart from CO2 fixation, the second largestsink for photosynthetic energy in manyhigher plants is nitrate. As the assimilationof nitrate in many species occurs predomi-nantly in the leaves, this process will oftenbe ongoing simultaneously with CO2 fixa-tion in photosynthetic cells (Noctor andFoyer, 1998).

Three processes related to photorespira-tion and nitrogen metabolism form impor-

tant links between chloroplasts, mitochon-dria, and peroxisomes. These are (1) gly-cine oxidation; (2) reductive amination ofoxoglutarate, and (3) hydroxypyruvate re-duction. These three processes are highlycoupled, and modulation of any one of themleads to a cascading effect on the other.

Glycine is formed in peroxisomes andoxidized in mitochondria. The precursor ofglycine is glycolate from chloroplasts. Gly-cine oxidation yields considerable amountsof NADH and NH4, besides CO2. The result-ing NADH is either used up for ATP produc-tion or exported out in the form of malate tomeet the requirements of hydroxypyruvatereduction in peroxisomes (Heldt et al., 1998;Raghavendra et al., 1998).

Oxidation of photorespiratory glycine iscoupled to hydroxypyruvate reduction(Hanning and Heldt, 1993; Heldt et al., 1998;Raghavendra et al., 1998). Any uncoupling ofglycine oxidation and hydroxypyruvate reduc-tion would imply a huge photorespiratory pro-duction of ATP, particularly in the mitochon-dria. If metabolic conditions demand, the extraNADH is used also for nitrate reduction incytosol. In algae, Weger et al. (1988) haveshown that high rates of NH4

+ assimilation areassociated with a marked increase in cyanide-sensitive O2 uptake.

The NADH generated by the glycinedecarboxylase reaction is expected to ac-count only for 50% of the reducing powerused in a subsequent reduction of thephotorespiratory hydroxypyruvate in per-oxisomes (Krömer and Heldt, 1991b). Theremainder of the reducing power is pro-vided by photosynthetic processes in thechloroplast (Krömer and Heldt, 1991b;Igamberdiev and Kleczkowski, 1997). Gly-cine oxidation can also increase theintramitochondrial and cytosolic ATP/ADPratio (Gardeström and Wigge, 1988); there-fore, the mitochondrial respiratory chain canplay a role in the cellular ATP production inthe light. Glycine oxidation in mitochondria

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of photosynthetic tissue in leaves of wheatand maize are coupled in more degree withcyanide-resistant and rotenone-resistantpaths of electron transport contrary to eti-olated leaves, where these pathways are in-volved to a much less extent (Igamberdievand Kleczkowski, 1997c; Igamberdiev etal., 1998).

Besides CO2, the other major sinks forreducing equivalents in chloroplasts are themetabolic reactions involving the reductionof nitrogen or sulfur. The reduction ofoxoglutarate to glutamate (or nitrite to NH4

+)occurs exclusively in chloroplasts. The majorsubstrate for these reactions, namely,oxoglutarate is exported from mitochondria,which operate a partial TCA cycle in light(see Section I.D). A continuous supply ofoxoglutarate from mitochondria is requiredfor NH4 assimilation into amino acids inchloroplasts. Similarly, the supply of nitriteto chloroplasts is also dependent on mito-chondrial activity, which provides signifi-cant amounts of NADH for nitrate reduc-tion in cytoplasm (Weger and Turpin, 1989;Padmasree and Raghavendra, 1998). Therequired NADH in mitochondria is gener-ated from glycine coming from peroxisomes.Thus, chloroplasts, mitochondria, and per-oxisomes have to work together to keep upthe reduction of nitrite and reductiveamination of oxoglutarate (Figure 10).

Glycine and malate, both of which areformed during active photosynthesis, form thesubstrates for leaf mitochondrial oxidation invivo. However, the main substrate for mito-chondrial respiration in the light is probablyglycine, which is produced at high rates duringphotorespiration. At least 25% of the NADHformed during oxidation of these metabolites isused for extra-mitochondrial requirements, par-ticularly hydroxypyruvate reduction in peroxi-somes and NO3– reduction in cytosol. The ex-port of reducing equivalents from mitochondriamay proceed by either a malate-aspartate shuttleor a malate-OAA shuttle. Chloroplasts form

alternative sources of reducing equivalents.Cytosolic nitrate reductase (NR) and peroxiso-mal hydroxypyruvate reductase can be servedvia a chloroplastic malate-OAA shuttle withreducing equivalents generated from photosyn-thetic electron transport (Heupel and Heldt,1992). Thus, mitochondrial metabolism be-comes a very important link among photosyn-thesis, photorespiration, and nitrogen assimila-tion in recycling NH4

+, reduced equivalents,and carbon skeletons (Figure 10).

F. Role in C 4 photosynthesis

Mitochondria play a direct role in car-bon metabolism of certain C4 and CAMplants, particularly those utilizing NAD-malic enzyme or PEP carboxykinase for C4-acid decarboxylation. In these plants thedecarboxylation of malate or aspartate oc-curs in mitochondria. During this function,mitochondria supply not only the carbonskeletons but also extra ATP needed for C4

pathway. The photosynthetic rates attainedby NAD+-malic enzyme plants suggest thatcarbon flux through the bundle sheath mito-chondria is 10- to 20-fold greater than thestandard respiratory carbon flux, andseveralfold greater than the flux of glycinethrough the mitochondria of C3 plants dur-ing photorespiration. Further, the NADHgenerated by NAD+-malic enzyme is uti-lized also for ATP synthesis. The predictedstoichiometry is about two malate moleculesoxidized per five molecules of PEP pro-duced (Siedow and Day, 2000). In PEPcarboxykinase type plants, the situation ismore complex.

The C4 plants do not show any photo-respiration because the process is confinedto bundle sheath cells and any CO2 releasedout is refixed efficiently in the surroundinglayer of mesophyll cells. Thus in C4 plants,mitochondrial location of GDC and result-

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FIG

UR

E 1

0. T

he i

nter

rela

tion

betw

een

nitr

ogen

met

abol

ism

, ch

loro

plas

t ph

otos

ynth

etic

rea

ctio

ns,

and

the

mito

chon

dria

lre

spira

tory

act

ivity

in p

lant

cel

ls. T

he in

itial

ste

p of

nitr

ate

redu

ctio

n to

nitr

ite o

ccur

s in

cyt

opla

sm. T

he m

ajor

ste

ps o

f for

mat

ion

and

assi

mila

tion

of a

mm

onia

are

loca

ted

in c

hlor

opla

sts.

The

red

ucin

g po

wer

for

nitr

ogen

ass

imila

tion

is s

uppl

ied

from

bot

hch

loro

plas

ts a

nd c

ytos

ol. T

he p

rovi

sion

of c

arbo

n sk

elet

ons

for a

mm

onia

ass

imila

tion

as w

ell a

s th

e re

cycl

ing

of p

hoto

resp

irato

ryam

mon

ia is

faci

litat

ed b

y m

itoch

ondr

ia. T

he k

ey e

nzym

es in

volv

ed in

thes

e pr

oces

ses

are

CS

, Citr

ate

synt

hase

; GD

C, G

lyci

nede

carb

oxyl

ase;

GO

GA

T,

Glu

tam

ate

oxog

luta

rate

am

ino

tran

sfer

ase/

Glu

tam

ate

synt

hase

; G

P,

Pho

spho

glyc

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dehy

de d

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drog

enas

e; G

S, G

luta

min

e sy

nthe

tase

; ID

H, I

soci

trat

e de

hydr

ogen

ase;

MD

H, M

alat

e de

hydr

ogen

ase;

ME

, Mal

ic e

nzym

e; N

iR,

Nitr

ite r

educ

tase

; N

R,

Nitr

ate

redu

ctas

e; P

DC

, P

yruv

ate

deca

rbox

ylas

e; P

EP

C,

Pho

spho

enol

pyru

vate

car

boxy

lase

; P

K,

Pyr

uvat

e ki

nase

.

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ing CO2 efflux becomes a crucial factor.Only the mitochondria located in bundlesheath cells of C4 palnts possess GDC, butnot those of mesophyll cells. The inter- andintracellular localization of GDC thus fa-cilitates the function of not only C4 photo-synthesis but also C3-C4 intermediacy (Deviet al., 1995; Rawsthorne, 1998).

V. BIOCHEMICAL BASIS:INTERORGANELLEINTERACTION

Rapid movement of metabolites occursbetween chloroplasts, cytoplasm, mitochondria,and peroxisomes. Such metabolite movementforms an important basis of interorganelle in-teraction as well as the optimization of differentmetabolic pathways in a plant cell. Severalinvestigators therefore attempted to study themetabolite patterns as the biochemical basis ofessentiality of mitochondrial respiration for pho-tosynthetic carbon assimilation, under variedconditions, for example, limiting or saturatingCO2, variable light intenstity (Krömer et al.,1988; Krömer et al., 1992; Krömer et al., 1993;Igamberdiev et al., 1997a,b, 1998; Padmasreeand Raghavendra, 1999a,b,c). These metabo-lites can be categorized into four groups:

1. Metabolites related to redox status, e.g.,malate or triose-P (mainly DHAP)

2. Adenylate compounds such as ATP orADP

3. Metabolites related to sucrose biosyn-thesis

4. Intermediates of the Calvin cycle

A. Major Products of OrganelleMetabolism

The interaction between the chloroplastsand mitochondria often involves not only

cytosol but also peroxisomes. Within thecell, there is always a high demand for ATPand reducing equivalents. The responsibil-ity of meeting the cellular requirements ofATP and NADH is shared by both chloro-plasts and mitochondria. The metabolismwithin chloroplasts, mitochodria, or peroxi-somes is optimized only when these or-ganelles are able to export their metabolitesand keep up the interorganelle metabolitemovement. The export of reduced equiva-lents from chloroplasts is essential to pre-vent overreduction of chloroplasts.

1. Chloroplasts

The major products exported from chlo-roplasts of C3 plants in light are glycolate,triose-P, and malate. The pattern of exportdepends on the carboxylase vs. oxygenaseactivity of Rubisco, which in turn dependson the ambient CO2. While the carboxylaseactivity of Rubisco results in the formationof triose-P, the oxygenase activity of Rubiscoleads to the formation of glycolate. Becausethe ratio of oxygenation to carboxylationduring photosynthesis in a leaf is 0.2 to 0.5,very high metabolic flux of glycolate oc-curs through the leaf peroxisomes (Heupelet al., 1991; Reumann et al., 1994). About50 to 75% of carbon from glycolate is sal-vaged in a sequence of photorespiratoryreactions that involves cooperation betweenthe chloroplasts, the mitochondria, and theperoxisomes.

Part of the triose-P formed by the reduc-tion of 3-PGA in chloroplasts is used up toregenerate RuBP, while the majority is ex-ported out to be utilized for either sucrosesynthesis or respiratory glycolytic pathway.Triose-phosphates are exported in exchangefor Pi through triose-P-Pi translocator lo-cated on the chloroplast inner membrane(Flügge, 1999). Triose-P exported mostly inthe form of DHAP to cytosol serves two

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major purposes: (1) form sucrose, (2) oxi-dation to PGA releasing ATP and NADH tomeet the needs of cytosol.

In light, the export of malate from chloro-plasts plays a significant role in the transfer ofreducing equivalents formed in excess of thoserequired to operate Calvin cycle. When theNADPH/NADP+ ratio in the chloroplast ishigh, OAA is converted to malate and ex-ported via the dicarboxylate translocator inthe inner envelope membrane of the chloro-plasts (Heineke et al., 1991). For the malate/OAA shuttle to operate as an effective NADPHexport system, the exported malate must beoxidized to regenerate OAA for transport backto the chloroplast. Thus, malate released intothe cytosol is either oxidized in cytosol tosupport nitrate reduction or transferred to per-oxisomes to support hydyroxypyruvate reduc-tion (Atkin et al., 2000b). Under conditionswhere more reductant is produced than is re-quired for cytosolic and peroxisomal processes,malate can be imported into mitochodria foroxidation and allow ATP synthesis (Hoefnagelet al., 1998).

Among the products of chloroplast me-tabolism, glycolate is the substrate forphotorespiratory metabolism, which helpsin the dissipation of excess energy as wellas protection against photoinhibition (seeSection III.C). Triose-P and malate facili-tate export of the reducing power and ATPfrom chloroplasts and thus act as sinks. Atlimiting CO2 malate is the major carrier ofreducing equivalents sent out of chloroplasts,while at optimal CO2, triose-P becomes thedominant carrier of reducing equivalents andleads to the formation of sucrose.

2. Peroxisomes

Glycine and glycerate are the majorproducts exported from peroxisomes. In theperoxisomes, glycolate is oxidized to

glyoxylate and then to glycine usingglutamate as the amino donor.

Glycine is exported from peroxisomesto mitochondria. On the other hand, peroxi-somes import serine from mitochondria andconvert it to hydroxypyruvate. The reduc-tion of hydroxypyruvate leads to the forma-tion of glycerate, which is exported to chlo-roplasts, facilitating the salvage of carbon.The reduction of hydroxypyruvate toglycerate places a high demand for reduc-ing equivalents. This demand is met in theform of malate exported to peroxisomes fromboth chloroplasts and mitochondria (Heldtet al., 1998; Raghavendra et al., 1998).

The metabolites within the peroxisomesare channelled through multienzyme com-plexes located in the matrix of peroxisomes.The metabolite movement into and out ofperoxisomes occurs through specific porescalled ‘porins’ (Reumann et al., 1995).

3. Mitochondria

The three major compounds exportedfrom mitochondria are serine (participatesin photorespiratory cycle), oxoglutarate (tosupply carbon compounds for N2 metabo-lism), and malate (to transfer reducingequivalents to peroxisomes).

The glycine formed in the peroxisomesis transported into the mitochondria, whereit is oxidized by glycine decarboxylase-serine hydroxymethyl transferase complexto yield serine, CO2, NH+

4, and NADH(Oliver, 1998; Douce and Neuberger, 1999).Serine leaves the mitochondria via a spe-cific translocator, possibly the sametranslocator by which glycine is taken up.

Carbon intermediates, particularly2-oxoglutarate, are exported from the TCAcycle to support GOGAT activity forglutamate synthesis in the chloroplasts (Fig-ure 10). While the major route of

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2-oxoglutarate production involves partialoperation of the TCA cycle in the mito-chondrion, 2-oxoglutarate synthesis mayalso occur via a cytosolic isocitrate dehy-drogenase (see Section I.D., Figure 2). Inmature leaves, the most important input ofC4 acids for 2-oxoglutarate synthesis ap-pears to be as oxaloacetate, generated bycytosolic phosphoenolpyruvate carboxylase.Approximately half of the PEP available incytosol is carboxylated by PEPC to oxalo-acetate, which is converted to 2-oxoglutaratethrough reactions catalyzed by citrate syn-thase, aconitase and isocitrate dehydroge-nase (Foyer et al., 2000).

In contrast to mitochondria from animaltissues, whose inner membrane is impermeableto oxaloacetate, the plant mitochondrial innerenvelope membrane has a malate-oxaloacetatetranslocator that facilitates the exchange ofmalate and oxaloacetate (Ebbighausen et al.,1985; Douce and Neuburger, 1990). The highactivity of malate dehydrogenase in the mito-chondrial matrix ensures an efficient reductionof oxaloacetate to malate. Thus, the NADHformed during glycine oxidation is incorpo-rated into malate and is exported by the malate-oxaloacetate shuttle. This shuttle has a highcapacity to transfer reducing equivalents frommitochondria. Although the amount of NADHgenerated in the mitochondria from glycineoxidation is quite high, mitochondria deliveronly about half the reducing equivalents re-quired for peroxisomal hydroxypyruvate re-duction, while the rest is provided by the chlo-roplasts (Heldt et al., 1998; Padmasree andRaghavendra, 2000).

B. Metabolite Exchange betweenChloroplasts, Mitochondria,Peroxisomes, and Cytosol

ATP and NAD(P)H are required in sev-eral steps of metabolic reaction occurring in

different cellular compartments. However,ATP and NAD(P)H being not permeableacross the membrane have to be transportedindirectly through different metaboliteshuttles. The rapid exchange of metabolitesbetween chloroplasts, mitochondria, peroxi-somes, and cytosol according to the cellularneeds of energy demand is the biochemicalbasis as well as essential component ofinterorganelle interaction (Figure 11).

During illumination, the difference inredox potentials between the stromal com-partment (NADPH/NADP) and cytosol(NADH/NAD) is quite large, leading to thetransfer of redox equivalents from the chlo-roplast stroma to the cytosol (Heineke et al.,1991). The transfer of reducing equivalentsfrom chloroplasts is mediated by two differ-ent metabolite shuttles: the triose-P-PGAshuttle mediated by the phosphate translocatorand the malate-OAA shuttle facilitated bythe dicarboxylate translocator. The triose-P-PGA shuttle is regulated by Pi availabilityfor counter-exchange by the phosphatetranslocator, as well as PGA reduction inchloroplasts and triose-P oxidation in cyto-sol. On the other hand, the malate-OAAshuttle is regulated by stromal NADP-MDHand the [NADPH]/[NADP], and also by thetranslocating step across the innerchloroplastenvelope membrane. In addition, metaboliteshuttles of malate and OAA between mito-chondria and the cytosol as well as cytosoland peroxisomes facilitate further the ex-change of reducing equivalents between mi-tochondria, cytosol, and peroxisomes (Heldt,1997).

A photosynthetic cell has two differentsystems to produce and meet cytosolic de-mands of ATP: photophosphorylation andoxidative phosphorylation. The ATP pro-duced during photophosphorylation is trans-ferred from chloroplast to cytosol throughthe exchange of triose-P and PGA mediatedby triose-P-Pi translocator. An NAD-depen-dent glyceraldehyde phosphate dehydroge-

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FIG

UR

E 1

1. T

he b

ioch

emic

al b

asis

of i

nter

orga

nelle

inte

ract

ion

betw

een

chlo

ropl

asts

, cyt

osol

, mito

chon

dria

, and

per

oxis

omes

. The

rapi

d m

ovem

ent

of m

etab

olite

s be

twee

n th

ese

orga

nelle

s fa

cilit

ates

the

exp

ort

of r

educ

ed e

quiv

alen

ts a

s w

ell

as A

TP

fro

m c

hlor

opla

sts

and

mito

chon

dria

.P

erox

isom

es f

orm

a m

ajor

sin

k fo

r re

duce

d eq

uiva

lent

s, w

hile

AT

P is

nee

ded

for

seve

ral a

ctiv

ities

(in

clud

ing

the

sucr

ose

bios

ynth

esis

) in

cyt

osol

.T

he m

etab

olite

shu

ttle

is fa

cilit

ated

by

spec

ific

carr

ier

prot

eins

on

the

mem

bran

es, c

alle

d tr

ansl

ocat

ors,

whi

ch a

re in

dica

ted

by n

umbe

rs 1

to 6

. The

glyc

olat

e/gl

ycer

ate

tran

sloc

ator

(1)

loc

ated

on

inne

r ch

loro

plas

t m

embr

ane

and

glyc

ine/

serin

e tr

ansl

ocat

or (

2) l

ocat

ed o

n in

ner

mito

chon

dria

lm

embr

ane

coor

dina

te th

e m

etab

olite

traf

fic in

volv

ing

maj

or p

hoto

resp

irato

ry m

etab

olite

s, b

etw

een

chlo

ropl

asts

, per

oxis

omes

, and

mito

chon

dria

. The

othe

r tw

o m

ajor

gat

eway

s in

volv

ed a

re th

e P

i tra

nslo

cato

r (2

) an

d di

carb

oxyl

ate

tran

sloc

ator

(3)

in c

hlor

opla

sts.

In m

itoch

ondr

ia, t

he o

ther

cha

nnel

sar

e th

e ad

enyl

ate

tran

sloc

ator

(5)

and

oxa

loac

etat

e tr

ansl

ocat

or (

6). T

he m

itoch

ondr

ial e

lect

ron

tran

spor

t (E

TC

) sy

stem

can

oxi

dize

ext

erna

l as

wel

las

inte

rnal

NA

DH

and

gen

erat

e A

TP

. (A

dapt

ed f

rom

Pad

mas

ree

and

Rag

have

ndra

, 19

98.)

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nase plays a significant role in facilitatingavailability of the chloroplastic ATP to cy-tosolic demands (Krömer, 1995). On theother hand, ATP produced during oxidativephosphorylation in mitochondria can betransferred directly to cytosol through ade-nylate translocator.

Illuminated chloroplasts usually haveexcess NADPH or related metabolites be-cause their electron transport activity is inmuch excess of the capacity of carbon fixa-tion (Huner et al., 1998). The excess reduc-ing equivalents are transported from the chlo-roplasts (in the form of DHAP and malate)to the cytosol to generate NADH. Mito-chondria are capable of oxidizing externalNADH. However, the oxidation could alsobe indirect through the shuttles of relatedmetabolites formed during photosynthesis(Gardeström et al., 2001). For example, gly-colate/glycerate translocator of chloroplastsand glycine/serine translocator of mitochon-dria can channel large amounts of glycineinto mitochondria. As glycine is the preferedmitochondrial substrate over malate, underphotorespiratory conditions NADH gener-ated during glycine oxidation can besuccesfully oxidized through the nonphos-phorylating pathways of mitochondrial elec-tron transport even when ADP is limited.Half of the reducing equivalents producedin the mitochondrial matrix are transferredto peroxisomes in the form of malate tosupport hydroxypyruvate reduction, whilethe rest is supplemented by malate derivedfrom chloroplasts.

On the basis of the metabolite movementsdescribed above, the photosynthetic and res-piratory activity in chloroplasts and mitochon-dria, respectively, appears to be modulated byone or more of the following factors: (1) theredox state due to the relative levels of NAD(P)or NAD(P)H (b) interorganelle movement ofmetabolites such as PGA, DHAP, malate, andOAA and (c) adenine nucleotides (ATP, ADP).Peroxisomes and cytoplasm naturally and

closely linked to these processes and form anactive and integral components of metabolitemovements and subsequent interorganelle in-teraction.

VI. FUTURE PERSPECTIVES

Most of the experiments on the interac-tion between mitochondria and chloroplastsduring photosynthesis have been made withprotoplasts (e.g., Gardeström et al., 1992;Krömer et al., 1988, 1993; Igamberdiev etal., 1998; Padmasree and Raghavendra,1999a,b,c). Only a few experiments wereconducted with intact leaves (Krömer andHeldt, 1991a; Hanson, 1992; Hanning andHeldt, 1993; Hurry et al., 1995; Atkin et al.,1998). However, more experiments areneeded using intact tissues or leaf discs soas to understand and extrapolate the situa-tion in leaves because the isolated proto-plasts lack the cell wall typical of plant cellsand may deviate in their metabolism.

An inherent limitation of metabolic in-hibitors is the possibility of their unspecificand multiple effects on different processes inthe cell. For example, SHAM, used exten-sively to inhibit alternative oxidase pathway,may also affect chlororespiration (Singh etal., 1992), chloroplastic glycolate-quinoneoxidoreductase (Goyal and Tolbert, 1996),besides stimulating peroxidase (Lambers,1985; Møller et al., 1988). Similarly, antimy-cin A used to suppress cytochrome pathwaymay also affect chlororespiration (Singh etal., 1992) as well as photosynthetic O2 evolu-tion (Cornic et al., 2000) due to the interfer-ence with ferredoxin-dependent reduction ofcyt b-559 particularly in intact chloroplasts(Scheller, 1996; Endo et al., 1998; Ivanov etal., 1998) besides stimulation of carbon fixa-tion (Schacter and Bassham, 1972). There-fore, the use of inhibitors to examine the roleand importance of the cytochrome and alter-

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native pathways has been questioned fre-quently. Neverthless, these metabolic inhibi-tors were used in mitochondrial studies bychoosing carefully the concentrations thataffect only mitochondrial respiration but notchloroplast reactions (Igamberdiev et al.,1997a,b; Padmasree and Raghavendra,1999a,b,c).

The respiratory measurements often uti-lize the Clark-type oxygen electrode, whichmonitors only the net changes in the O2

levels (caused by both consumption of O2 inrespiration and evolution of O2 in photosyn-thesis). It is desirable that these two pro-cesses be monitored seperately, so as tomake precise measurements of photosyn-thesis or respiration. Mass spectrophotom-eter, which can monitor 18/16O2 or 13/12CO2,has been extremely useful for not only dis-tinguishing between photosynthesis and res-piration (Avelange et al., 1991) but also tomake quantitative measurements of alterna-tive pathway activity (Robinson et al., 1995).

The studies using inhibitors can becomplemented by experiments involvingmutants or transgenic plants, with an alteredpattern of proteins/enzymes related to chlo-roplasts, mitochondria, and peroxisomes.Extensive studies are made on transgenicplants with overexpression or (antisense)depression of enzymes such as triose-P de-hydrogenase, rubisco, activase, rubisco orproteins such as triose-P-phosphate-translocator (Vivekanandan and Saralabai,1997; Heineke, 1998; Sharkey, 1998; Huber,1998; Flügge, 2000; Häusler et al., 2000).Similarly, mutants or transgenics with al-tered levels of invertase or sucrose synthaseor ADP-glucose pyrophosphorylase or PRKor FBPase or glutamine synthetase orglutamate synthase are available (Häusler etal., 1994; Heineke, 1998; Paul et al., 2000).In contrast, only a few studies are made onmutants/transgenics with altered respiratorycharacteristics (Vanlerberghe et al., 1994;Hiser et al., 1996; Gutierres et al., 1997;

Igamberdiev et al., 2001). In an interestingrecent study, Sabar et al. (2000) used themale sterile mutants of Nicotiana sylvestristo study some aspects of respiration andphotosynthesis. So far, no studies have beenreported on the chloroplast-mitochondriainteractions in suitable transgenic plants.

The plant mitochondria have an uniquesystem of two different types of oxidativeelectron transport-cytochrome pathway andthe alternative pathway. Being a major routefor ATP formation in mitochondria, theimportance of cytochrome pathway is un-questionable and is obvious. However, thephysiological importance of alternative path-way is not completely understood. Oxida-tive electron transport in mitochondria oc-curs predominantly through alternativepathway during glycine oxidation in mito-chondria or LEDR in barley protoplasts(Igamberdiev et al., 1997a,b). This phenom-enon has to be analyzed further preferablyby employing tools other than the metabolicinhibitors. Further, the role of alternativepathway in optimizing chloroplast functionalso has to be studied under varied environ-mental conditions, such as light intensity,temperature, and stress conditions. The re-sults would be quite exciting and wouldhelp us understand not only interorganelleinteraction but also the alternative pathwayitself, which is unique to plant mitochon-dria.

The strong interaction between chlo-roplasts, mitochondria, peroxisomes, andcytosol is possible only when there is aneffecient cross-talk between these or-ganelles. Obviously, metabolite move-ment is an important factor or signal.However, it is possible that there are othersignals. Further work is necessary to iden-tify and establish the importance of dif-ferent signals between the organelles.Among the possibilities are cytosolic pH,phosphate status, and even the superox-ide radicals.

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The major advantage of the interorganelleinteraction appears to be optimization of theirfunction and protection from any damagedue to the unfavorable factors. For example,the chloroplasts are protected from gettingoverreduced. It is quite likely that mitochon-dria are also prevented from gettingoveroxidized. However, not much informa-tion is available pertaining to protection ofmitochondria.

On exposure to supra-optimal light andlikely photoinhibition, the mitochondria(along with peroxisomes) rescue the chloro-plasts by dissipating their excess reducedequivalents (Saradadevi and Raghavendra,1992; Hurry et al., 1998; Padmasree andRaghavendra, 2000; Gardeström et al.,2001). It would be of great interest andexciting to examine the interaction betweenthe different organelles, particularly chloro-plasts, mitochondria, and peroxisomes andthe consequences on metabolic regulationwhen the plant is subjected to other stressconditions such as temperature or water.

ACKNOWLEDGMENTS

Work in our laboratory on photosynthe-sis and respiration in mesophyll and guardcell protoplasts was supported by a grant(No. SP/SO/A-12/98) from Department ofScience and Technology, New Delhi toA.S.R. K.P. is a recipient of ResearchAssociateship and L.P. holds a Senior Re-search Fellowship, both from the Council ofScientific and Industrial Research, New Delhi.

REFERENCES

Anderson, J.M. 2001. Strategies of photosyn-thetic adaptations and acclimation. In :Probing Photosynthesis. Mechanisms,

Regulation and Adaptation. pp. 283–292.Yunus, M., Pathre, U., and Mohanty, P.,Eds., Taylor and Francis Press, Londonand New York.

Andersson, B. and Barber, J. 1996. Mechanismsof photodamage and protein degradationduring photoinhibition of photosystem II.In : Photosynthesis and the Environment.pp. 101–121. Baker, N.R., Ed., KluwerAcademic, Dordrecht.

Atkin, O.K., Cummins, W.R. and Collier, D.E.1993. Light induction of alternative path-way capacity in leaf slices of Belgiumendive. Plant Cell Environ. 16:231–235.

Atkin, O.K., Evans, J.R., Ball, M.C., Lambers,H. and Pons, T.L. 2000a. Leaf respirationof snowgum in light and dark. Interactionsbetween temperature and irradiance. PlantPhysiol. 122:915–923.

Atkin, O.K., Millar, A.H., Gardeström, P., andDay, D.A. 2000b. Photosynthesis, carbo-hydrate metabolism and respiration inleaves of higher plants. In : Photosynthe-sis: Physiology and Metabolism. pp. 153–175. Leegood, R.C., Sharkey, T.D. andvon Caemmerrer, S., Eds., Kluwer Aca-demic Publishers, The Netherlands.

Atkin, O.K., Seibke, K., and Evans, J.R. 1998.Relationship between the inhibition of leafrespiration by light and enhancement ofleaf dark respiration following light treat-ment. Aus. J. Plant Physiol. 25:437–443.

Atkin, O.K., Westbeek, M.H.M., Cambridge,M.L., Lambers, H., and Pons, T.L. 1997.Leaf respiration in light and darkness. Acomparison of slow and fast growing Poaspecies. Plant Physiol. 113:961–965.

Avelange, M.H., Thiéry, J.M., Sarrey, F., Gans,P., and Rébeillé, F. 1991. Mass spectro-metric determination of O2 and CO2 gasexchange in illuminated higher plant cells.Evidence for light inhibition of substratedecarboxylations. Planta 183:150–157.

Azcón-Bieto, J. 1992. Relationships betweenphotosynthesis and respiration in the dark

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nly.

111

in plants. In: Trends in PhotosynthesisResearch. pp. 241–253. Barber, J.,Guerrero, M.G., and Medrano, H., Eds.,Intercept, Andover.

Backhausen, J.E., Kitzmann, C., and Scheibe, R.1994. Competition between electron accep-tors in photosynthesis: Regulation of themalate valve during CO2 fixation and ni-trite reduction. Photosynth. Res. 42:75–86.

Budde, R.J.A. and Randall, D.D. 1990. Pea leafmitochondrial pyruvate dehydrogenasecomplex is inactivated in vivo in a lightdependent manner. Proc. Natl. Aca. Sci.USA. 87:673–676.

Carpentier, R. 1997. Influence of high light onphotosynthesis: photoinhibition and energydissipation. In : Handbook of Photosyn-thesis. pp. 443–450. Pessarakli, M., Ed.,Marcel Dekker, New York.

Ceulemans, R. and Saugier, B. 1991. Photosyn-thesis. In: Physiology of Trees. pp. 21–50.Raghavendra, A.S., Ed., John Wiley, NewYork.

Chen, R.C. and Gadal, P. 1990. Do the mito-chondria provide the 2–oxogluterateneeded for glutamate synthesis in higherplant chloroplasts? Plant Physiol. Biochem.28:141–145.

Cornic, G., Bukhov, N.G., Wiese, C., Bligny, R.and Heber, U. 2000. Flexible couplingbetween light-dependent electron and vec-torial proton transport in illuminated leavesof C3 plants. Role of photosystem I-de-pendent proton pumping. Planta 210:468–477.

Critchley, C. 1998. Photoinhibition. In : Photo-synthesis: A Comprehensive Treatise. pp.264–272. Raghavendra, A.S., Ed., Cam-bridge University Press, Cambridge.

Devi, T., Rajagopalan, A.V. and Raghavendra,A.S. 1995. Predominant localization of mi-tochondria enriched with glycine-decarboxy-lating enzymes in bundle sheath cells ofAlternanthera tenella, a C3–C4 intermediatespecies. Plant Cell Environ. 18:589–594.

Diethelm, R., Miller, M.G., Shibles, R., and Stewart,C.R. 1990. Effect of salicylhydroxamic acidon respiration, photosynthesis and peroxidaseactivity in various plant tissues. Plant CellPhysiol. 31:179–185.

Douce, R. and Neuburger, M. 1990. Metaboliteexchange between the mitochondrion andthe cytosol. In : Plant Physiology, Biochem-istry and Molecular Biology. pp. 173–190.Dennis, D.T. and Turpin, D.H., Eds.,Longman Scientific & Technical, Harlow.

Douce, R. and Neuburger, M. 1999. Biochemi-cal dissection of photorespiration. Curr.Opinion Plant Biol. 2:214–222.

Ebbighausen, H., Chen, J. and Heldt, H.W. 1985.Oxaloacetate translocator in plant mitochon-dria. Biochem. Biophys. Acta 810:184–199.

Edwards, G. and Walker, D.A. 1983. C3, C4:Mechanisms, and Cellular and Environ-mental Regulation, of Photosynthesis,Blackwell Scientific Publications, Oxford.

Ekelund, N.G. 2000. Interactions between pho-tosynthesis and light enhanced dark respi-ration (LEDR) in the flagellate Euglenagracilis after irradiation with ultravioletradiation. J. Photochem. Photobiol. 55:63–69.

Endo, T., Shikanai, T., Sato, F., and Asada, K.1998. NAD(P)H dehydrogenase-depen-dent, antimycin A-sensitive electron do-nation to plastoquinone in tobacco chloro-plasts. Plant Cell Physiol. 39:1226–1231.

Faske, M., Holtgrefe, S., Ocheretina, O., Meister,M., Backhausen, J.E. and Scheibe, R. 1995.Redox equilibria between the regulatorythiols of light/dark-modulated chloroplastenzymes and dithiothreitol: fine-tuning bymetabolites. Biochim. Biophys. Acta1247:135–142.

Flügge, U-I. 1999. Phosphate translocators inplastids. Annu. Rev. Plant Physiol. PlantMol. Biol. 50:27–45

Flügge, U-L. 2000. Metabolite transport acrossthe chloroplast envelope of C3 plants.

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

112

In : Photosynthesis: Physiology and Me-tabolism. pp. 137–152. Leegood, R.C.,Sharkey, T.D. and von Caemmerer, S.,Eds., Kluwer Academic Publishers,Dordrecht.

Flügge, U-I. and Heldt, H.W. 1991. Metabolitetranslocators of the chloroplast envelope.Annu Rev. Plant Physiol. Plant Mol. Biol.42:129–144.

Foyer, C.H., Ferrario-Méry, S., and Huber, S.C.2000. Regulation of carbon fluxes in thecytosol: Coordination of sucrose synthe-sis, nitrate reduction and organic acid andamino acid biosynthesis. In: Advances inPhotosynthesis, Photosynthesis: Physiol-ogy and Metabolism. Vol. 9. pp. 177–203.Leegood, R.C., Sharkey, T.D. and vonCaemmerer, S., Eds., Kluwer AcademicPublishers, Dordrecht.

Gans, P. and Rébeillé, F. 1988. Light inhibitionof mitochondrial respiration in a mutant ofChlamydomonas reinhardtii devoid ofribulose-1,5–bisphosphate carboxylase/oxygenase activity. Arch. Biochem.Biophys. 260:109–117.

Gardeström, P. 1996. Interactions between mi-tochondria and chloroplasts. Biochim.Biophys. Acta 1275:38–40.

Gardeström, P. and Lernmark, U. 1995. Thecontribution of mitochondria to energeticmetabolism in photosynthetic cells. J.Bioenerg. Biomembr. 27:415– 421.

Gardeström, P. and Wigge, B. 1988. Influenceof photorespiration on ATP/ADP ratios inthe chloroplasts, mitochondria and cyto-sol, studied by rapid fractionation of bar-ley (Hordeum vulgare) protoplasts. PlantPhysiol. 88:69–76.

Gardeström, P., Bergman, A., and Ericson, I.1981. Inhibition of the conversion of gly-cine to serine in spinach leaf mitochon-dria. Physiol. Plant. 53:439–444.

Gardeström, P., Igamberdiev, A.U., andRaghavendra, A.S. 2002. Mitochondrialfunctions in light. In :–Photosynthetic Ni-

trogen Assimilation and Associated Car-bon Metabolism. Foyer, C. and Noctor,G., Eds., Kluwer Academic Publishers,Dordrecht. in press.

Gardeström, P., Zhou, G., and Malmberg, G.1992. Respiration in barley protoplastsbefore and after illumination. In : Molecu-lar, Biochemical and Physiological As-pects of Plant Respiration. pp. 261–265.Lambers, H. and van der plas, L.H.W.,Eds., S.P.B Academic Publishing, TheHague.

Gautier, A., Vavasseur, A., Gans, P., and Lascéve,G. 1991. Relationship between respirationand photosynthesis in guard cell and meso-phyll protoplasts of Commelina communisL. Plant Physiol. 95:636–641.

Gillmore, M. 1997. Mechanistic aspects of xan-thophyll cycle-dependent photoprotectionin higher plant chloroplasts and leaves.Physiol. Plant. 99:197–209.

Gonzàlez-Meler, M.A., Ribas-Carbo, M., Giles,L. and Siedow, J. 1999. The effect ofgrowth and measurement temperature onthe activity of the alternative pathway.Plant Physiol. 120:765–772.

Gout, E., Bligny, R., Pascal, N., and Douce, R.1993. 13C nuclear magnetic resonance stud-ies of malate and citrate synthesis andcompartmentation in higher plant cells. J.Biol. Chem. 268:3986–3992.

Goyal, A. and Tolbert, N.E. 1996. Associationof glycolate oxidation with photosyntheticelectron transport in plant and algal chlo-roplasts. Proc. Natl. Acad. Sci. USA93:3319–3324.

Graham, D. 1980. Effects of light on dark res-piration. In : The Biochemistry of Plants. AComprehensive Treatise. pp. 525–579.Vol. 2., Davies, D.D., Ed., Academic Press,New York.

Graham, D. and Walker, D.A. 1962. Some ef-fects of light on the inter conversion ofmetabolites in green leaves. Biochem. J.82:554–561.

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

113

Guo, L-W., Xu, D-Q., and Shen, Y-K. 1995.Protective effect of photorespirationaganist photoinhibition in cotton leaves.In : Photosynthesis: from Light to Bio-sphere. pp.195–198. Vol. 4., Mathis, P.,Ed., Kluwer Academic, Dordrecht.

Gutierres, S., Sabar, M., Lelandais, C., Chetrit,P., Diolez, P., Degand, H., Boutry, M.,Vedel, F., de Kouchkovsky, Y., and DePaepe, R. 1997. Lack of mitochondrialand nuclear encoded subunits of complexI and alteration of respiratory chain inNicotiana sylvestris mitochondrial dele-tion mutants. Proc. Natl.Acad. Sci. USA94:3436–3441.

Hanning, I. and Heldt, H.W. 1993. On the func-tion of mitochondrial metabolism duringphotosynthesis in spinach (Spinaciaoleracea L) leaves. Partitioning betweenrespiration and export of redox equiva-lents and precursors for nitrate assimila-tion products. Plant Physiol. 103:1147–1154.

Hanson, K.R. 1992. Evidence for mitochondrialregulation of photosynthesis by a starchlessmutant of Nicotiana sylvestris. PlantPhysiol. 99:276–283.

Häusler, R.E., Lea, P.J., and Leegood, R.C. 1994.Control of photosynthesis in barley leaveswith reduced actvities of glutamine syn-thetase or glutamate synthetase. Planta194:418–434.

Häusler, R.E., Schlieben, N.H., Nicolay, P.,Fischer, K., Fischer, K.L., and Flugge U-I. 2000. Control of carbon partitioning andphotosynthesis by the triose phosphate/phosphate translocator in transgenic to-bacco plant (Nicotiana tabaccum). I. Com-parative physiological analysis of tobaccoplants with antisense repression andoverexpression of the triose phosphate/phosphate translocator. Planta 210:371–382.

Heber, U., Bligny, R., Streb, R., and Douce, R.1996. Photorespiration is essential for theprotection of photosynthetic apparatus of

C3 plants aganist photoinactivation undersunlight. Bot. Acta. 109:307–315.

Heichel, G.H. 1971. Response of respiration oftobacco leaves in light and darkness andthe CO2 compensation concentration toprior illumination and oxygen. PlantPhysiol. 48:178–182.

Heineke, D. 1998. Application in biotechnol-ogy. In : Photosynthesis: A Comprehen-sive Treatise. pp. 352–361, Raghavendra,A.S., Ed., Cambridge University Press,Cambridge.

Heineke, D., Riens, B., Grosee, H., Hoferichter,P., Peter U., Flugge, U-I., and Heldt, H.W.1991. Redox transfer across the inner chlo-roplast envelope membrane. Plant Physiol.95:1131–1137.

Heldt, H.W. 1997. Plant Biochemistry andMolecular Biology, Oxford UniversityPress, Oxford.

Heldt, H.W., Raghavendra, A.S., Reumann, S.,Bettermann, M., Hanning, I., Benz, R.,and Maier, E. 1998. Redox transfer be-tween mitochodria and peroxisomes inphotorespiration. In: Plant Mitochondria:From Gene to Function. pp. 543–549,Møller, I.M., Gardeström, P., Glimelius,K. and Glaser, E., Eds., Backhuys Pub-lishers, The Netherlands.

Heupel, R. and Heldt, H.W. 1992. Redox trans-fer between mitochondria and peroxi-somes. In: Molecular, Biochemical andPhysiological Aspects of Plant Respira-tion. pp. 243–247, Lambers, H. and vander Plas, L.H.W., Eds., SPB Academic,The Hague

Heupel, R., Markgraf, T., Robinson, D.G., andHeldt, H.W. 1991. Compartmentation stud-ies on spinach leaf peroxisomes. Evidencefor channeling of photorespiratory metabo-lites in peroxisomes devoid of intact bound-ary membrane. Plant Physiol. 96:971–979.

Hill, S.A. and Bryce, J.H. 1992. Malate inhibi-tion and light enhanced dark respiration in

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

114

barley protoplasts. In : Molecular, Bio-chemical and Physiological Aspects ofPlant Respiration. pp. 221–230, Lambers,H. and van der plas, L.H.W., Eds., S.P.B.Academic Publishing, The Hague.

Hiser, C., Kapranov, P., and McIntosh, L. 1996.Genetic modification of respiratory capac-ity in potato. Plant Physiol. 110:277–286.

Hoefnagel, M.H.N., Atkin, O.K., and Wiskich, J.T.1998. Interdependence between chloroplastsand mitochondria in the light and the dark.Biochem. Biophys. Acta. 1366:235–255.

Huber, S.C. 1998. Starch-sucrose metabolismand assimilate partitioning. In : Photosyn-thesis: A Comprehensive Treatise. pp. 163–172, Raghavendra, A.S., Ed., CambridgeUniversity Press, Cambridge.

Huber, S.C. and Huber, J.L. 1996. Sucrose-phos-phate synthase. Annu. Rev. Plant Physiol.Plant Mol. Biol. 47:431–444.

Huner, P.A.N., Öquist, G., and Sarhan, F. 1998.Energy balance and acclimation to lightand cold. Trends Plant Sci. 3:224–230.

Hurry, V., Huner, N., Selstam, E., Gardeström,P., and Öquist, G. 1998. Photosynthesis atlow growth temperatures. In : Photosyn-thesis: A Comprehensive Treatise. pp. 238–249. Raghavendra, A.S., Ed., CambridgeUniversity Press, Cambridge.

Hurry, V., Tobiaeson, M., Krömer, S., Gardeström,P., and Öquist, G. 1995. Mitochondria con-tribute to increased photosynthetic capacityof leaves of winter rye (Secale cereale) fol-lowing cold hardening. Plant Cell Environ.18:69–76.

Hurry, V.M. and Huner, N.P.A. 1992. Effect ofcold hardening on sensitivity of winterand spring wheat leaves to short-termphotoinhibition and recovery of photosyn-thesis. Plant Physiol. 100:1283–1290.

Igamberdiev, A.U., Zhou, G., Malmberg, G.,and Gardeström, P. 1997a. Respiration ofbarley protoplasts before and after illumi-nation. Physiol. Plant. 99:15–22.

Igamberdiev, A.U., Bykova, N.V., and Gardeström,P. 1997b. Involvement of cyanide-resistentand rotenone-insensitive pathways of mito-chondrial electron transport during oxida-tion of glycine in higher plants. FEBS Lett.412:265–269.

Igamberdiev, A.U. and Kleczkowsi, L.A. 1997.Glyoxylate metabolism during photores-piration-A cytosol connection. In: Hand-book of Photosynthesis. pp. 269–279,Pessarakli, M., Ed., Marcel Dekker, Inc,New York.

Igamberdiev, A.U., Bykova, N.V., Lea, P.J.,and Gardeström, P. 2001. The role of pho-torespiration in redox and energy balanceof photosynthetic plant cells: a study withbarley mutant deficient in glycine decar-boxylase. Physiol. Plant. 111:427–438.

Igamberdiev, A.U., Hurry, V., Krömer, S., andGardeström, P. 1998. The role of mito-chondrial electron transport during photo-synthetic induction. A study with barley(Hordeum vulgare) protoplasts incubatedwith rotenone and oligomycin. Physiol.Plant. 104:431–439.

Ivanov, B., Kobayashi, Y., Bukhov, N.G., andHeber, U. 1998. Photosystem I-dependentcyclic electron flow in intact spinach chlo-roplasts: occurrence, dependence on re-dox conditions and electron acceptors andinhibition by antimycin A. Photosynth.Res. 57:61–67.

Kirshbaum, M.U.F. and Furquhar, G.D. 1987.Investigation of the CO2 dependence ofquantum yield and respiration in Eucalyp-tus pauciflora. Plant Physiol. 83:1032–1036.

Kozaki, A. and Takeba, G. 1996. Photorespira-tion protects C3 plants from photooxida-tion. Nature 384:557–560.

Krömer, S. 1995. Respiration during photosyn-thesis. Annu. Rev. Plant Physiol. PlantMol. Biol. 46:45–70.

Krömer, S. and Heldt, H.W. 1991a. On the roleof mitochondrial oxidative phosphoryla-

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

115

tion in photosynthesis metabolism as stud-ied by the effect of oligomycin on photo-synthesis in protoplasts and leaves of bar-ley (Hordeum vulgare). Plant Physiol.95:1270–1276.

Krömer, S. and Heldt, H.W. 1991b. Respirationof pea leaf mitochondria and redox trans-fer between the mitochondrial andextramitochodrial compartment. Biochim.Biophys. Acta 1057:42–50.

Krömer, S., Malmberg, G., and Gardeström, P.1992. Mitochondrial contribution to pho-tosynthetic metabolism at different lightintensities and CO2 concentrations in bar-ley leaf protoplasts. In: Research in Pho-tosynthesis. Vol. III. pp. 709–712, Murata,N., ed., Kluwer Academic Publishers,Netherlands.

Krömer, S., Malmberg, G., and Gardeström, P.1993. Mitochondrial contribution to pho-tosynthetic metabolism. A study with bar-ley (Hordeum vulgare) leaf protoplasts atdifferent light intensities and CO2 concen-trations. Plant Physiol. 102:947–955.

Krömer, S., Stitt, M., and Heldt, H. W. 1988.Mitochondrial oxidative phosphorylationparticipating in photosynthetic metabolismof a leaf cell. FEBS Lett. 226:352–356.

Lambers, H. 1982. Cyanide-respiration: a non-phosphorylating electron transport path-way acting as an energy overflow. Physiol.Plant. 55:478–485.

Lambers, H. 1985. Respiration in intact plantsand tissues: its regulation and dependenceon environmental factors, metabolism andinvaded organisms. In: Encyclopaedia ofPlant Physiology, New Series, HigherPlant Cell Respiration. Vol.18. pp. 418–473, Douce, R. and Day, D.A., Eds.,Springer-Verlag, Berlin.

Lemaire, C., Wollman, F-A., and Bennoun, P.1988. Restoration of phototropic growthin a mutant of Chlamydomonas reinhardtiiin which the chloroplast atpB gene of theATP synthase has a deletion: an example

of mitochondria dependent photosynthe-sis. Proc. Natl. Acad. Sci. USA. 85:1344–1348.

Leuthy, M.M., Miernyk, J.A., David, N.R., andRandall, D.D. 1996. Plant pyruvate dehy-drogenase complexes. In : Alpha-Keto AcidDehydrogenase Complexes. pp. 71–92.Patel, M.S., Roch, T.E., and Haris, R.A.,Eds., Birkhäuser Verlag Press, Basel.

Long. S.P., Humphries, S. and Falkowski, P.G.1994. Photoinhibition of photosynthesisin nature. Ann. Rev. Plant Physiol. PlantMol. Biol. 45:633–662.

Lynnes, J.A. and Weger, H.G. 1996. Azide-stimulated oxygen consumption by thegreen alga Selenastrum minutum. Physiol.Plant. 97:132–138.

Mackenzie, S. and McIntosh, L. 1999. Higherplant mitochondria. Plant Cell 11:571–585.

Mangat, B.S., Levin, W.B. and Bidwell, R.G.S.1974. The extent of dark respiration inilluminated leaves and its control by ATPlevels. Can. J. Bot. 52:673–681.

Mawson, B.T. 1993. Modulation of photosyn-thesis and respiration in guard and meso-phyll cell protoplasts by oxygen concen-tration. Plant Cell Environ. 16:207–214.

McCashin, B.G., Cossins, E.A., and Canvin,D.T. 1988. Dark respiration duringphotosynthesisin wheat leaf slices. PlantPhysiol. 87:155–161.

McIntosh, L. 1994. Molecular biology of thealternative oxidase. Plant Physiol. 105:781–786.

Millar, H. and Day, D.A. 1997. Alternative so-lutions to radical problems. Trends PlantSci. 2:289–290.

Møller, I.M., Bérczi, A., van der Plas, L.H.W.,and Lambers, H. 1988. Measurement ofthe activity and capacity of the alternativepathway in intact plant tissues: identifica-tion of problems and possible solutions.Physiol. Plant. 72:642–649.

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

116

Niyogi, K.K, Grossman, A.R., and Bjorkman,O. 1998. Arabidopsis mutants define a cen-tral role for the xanthophyll cycle in theregulation of photosynthetic energy con-version. Plant Cell 10:1121–1134.

Niyogi. K.K. 1999. Photoprotection revisited:genetic and molecular approaches. Ann.Rev. Plant Physiol. Plant Mol. Biol.50:333–359.

Noctor, G. and Foyer, C.H. 1998. A reevalua-tion of the ATP: NADPH budget duringC3 photosynthesis: a contribution fromnitrate assimilation and its associated res-piratory activity? J. Exp. Bot. 49:1895–1908.

Ohad, I., Sonoike, K., and Andersson, B. 2000.Photoinactivation of the two photosystemsin oxygenic photosynthesis: mechanismsand regulations. In : Probing Photosynthe-sis. Mechanisms, Regulation and Adapta-tion. pp. 293–309, Yunus, M., Pathre, U.and Mohanty, P., Eds., Taylor and Francis,London.

Oliver, D.J. 1998. Photorespiration and the C2

cycle. In: Photosynthesis: A Comprehen-sive Treatise. pp. 173–182, Raghavendra,A.S., Ed., Cambridge University Press,Cambridge.

Osmond, B., Badger, M., Maxwell, K., Bjorkman,O., and Legood, R. 1997. Too many pho-tons: photorespiration, photoinhibition andphotooxidation. Trends in Plant Sci. 2:119–121.

Padmasree, K. and Raghavendra, A.S. 1998. Inter-action with respiration and nitrogen metabo-lism. In : Photosynthesis: A ComprehensiveTreatise. pp. 197–211, Raghavendra, A.S.,Ed., Cambridge University Press, Cam-bridge.

Padmasree, K. and Raghavendra, A.S. 1999a.Importance of oxidative electron transportover oxidative phosphorylation in optimiz-ing photosynthesis in mesophyll proto-plasts of pea (Pisum sativum L.). Physiol.Plant. 105:546–553.

Padmasree, K. and Raghavendra, A.S. 1999b.Prolongation of photosynthetic inductionas a consequence of interference withmitochondrial oxidative metabolism inmesophyll protoplasts of pea (Pisumsativum L.). Plant Sci. 142:29–36.

Padmasree, K. and Raghavendra, A.S. 1999c.Response of photosynthetic carbon assimi-lation in mesophyll protoplasts to restictionon mitochondrial oxidative metabolism:Metabolites related to the redox status andsucrose biosynthesis. Photosynth. Res.62:231–239.

Padmasree, K. and Raghavendra, A.S. 2000.Photorespiration and interaction betweenchloroplasts, mitochondria and peroxi-somes. In : Probing photosynthesis:Mechanism, Regulation and Adaptation.pp. 245–261, Yunus, M., Pathre, U., andMohanty, P., Eds., Taylor and Francis,London.

Padmasree, K. and Raghavendra, A.S. 2001a.Consequences of restricted mitochindrialoxidative metabolism on photosyntheticcarbon assimilation in mesophyll proto-plasts: decrease in light activation of fourchloroplastic enzymes. Physiol. Plant.112:582–588.

Padmasree, K. and Raghavendra, A.S. 2001b.Restriction of mitochondrial oxidativemetabolism leads to suppression of photo-synthetic carbon assimilation but not ofphotochemical electron transport in peamesophyll protoplasts. Current Sci. 81:680–684

Padmavathi, L. and Raghavendra, A.S. 2001.Importance of cytochrome pathway ofmitochondrial electron transport over thealternative pathway during the Kok effectin leaf discs of pea (Pisum sativum).Physiol. Plant. 113: 430–434

Park, Y-I., Anderson, J.M., and Chow, W.S. 1996.Light inactivation of photosystem II andD1 protein synthesis in vivo is independentof the modulation of the photosynthetic

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

117

apparatus by growth irradiance. Planta198:300–309.

Parnik, T. and Keerberg, O. 1995. Decarboxyla-tion of primary and end products of pho-tosynthesis at different oxygen concentra-tions. J. Exp. Bot. 46:1439–1447.

Parvathi, K. and Raghavendra, A.S. 1995. Bioen-ergetic processes in guard cells related tostomatal function. Physiol. Plant. 93:146–154.

Paul, M.J., Driscoll, S.P., Andralojc, P.J., Knight.J.S., Gray, J.C., and Lawlor, D.W. 2000.Decrease of phosphoribulokinase activityby antisense RNA in transgenic tobacco:definition of the light environment underwhich phosphoribulokinase is not in largeexcess. Planta 211:112–119.

Raghavendra, A.S. and Vani, T. 1989. Respira-tion in guard cells: pattern and possiblerole in stomatal function. J. Plant Physiol.135:3–8.

Raghavendra, A.S., Padmasree, K., andSaradadevi, K. 1994. Interdependence ofphotosynthesis and respiration in plantcells: interactions between chloroplasts andmitochondria. Plant Sci. 97:1–14.

Raghavendra, A.S., Reumann, S. and Heldt,H.W. 1998. Participation of mitochondrialmetabolism in photorespiration. Reconsti-tuted system of peroxisomes and mito-chondria from spinach leaves. PlantPhysiol. 116:1333–1337.

Randall, D.D., Miernyk, J.A., David, N.R., Gemel,G., and Leuthy, M.H. 1996. Regulation ofleaf mitochondrial pyruvate dehydrogenasecomplex activity by reversible phosphory-lation. In : Protein Phosphorylation inPlants. pp. 87–103. Shewry, P.R., Halford,N.G., and Hooley., Eds., Clarendon Press,Oxford.

Rawsthorne, S and Bauwe, H. 1998. C3–C4 in-termediate photosynthesis. In : Photosyn-thesis: A Comprehensive Treatise. pp. 150–162, Raghavendra, A.S., Ed., CambridgeUniversity Press, Cambridge.

Reddy, M.M., Vani, T., and Raghavendra, A.S.1991. Light enhanced dark respiration inmesophyll protoplasts from leaves of pea.Plant Physiol. 96:1368–1371.

Reumann, S., Heupel, R., and Heldt, H.W. 1994.Compartmentation studies on spinach leafperoxisomes. II. Evidence for the transferof reductant from the cytosol to peroxiso-mal compartment via malate-oxaloacetateshuttle. Planta 193:167–173.

Reumann, S., Maier, E., Benz, R., and Heldt,H.W. 1995. The membrane of leaf peroxi-somes contains a porin-like channel.J.Biol.Chem. 270:17559–17565.

Ribas-Carbo, M., Aroca, R., Gonzàlez-Meler,M.A., Irigoyen, J.J., and Sanchez-Diaz,M. 2000. The electron partitioning betweenthe cytochrome and alternative respiratorypathways during chilling recovery in twocultivars of maize differing in chillingsensitivity. Plant Physiol. 122:199–204.

Robinson, S.A., Ribas-Carbo, M., Yakir, D.,Giles, L., Reuveni, Y., and Berry, J.A.1995. Beyond SHAM and cyanide-oppor-tunities for studying the alternative oxi-dase in plant respiration using oxygen iso-tope discrimination. Aust. J. Plant Physiol.22: 487–496.

Rychter, A.M., Ciesla, E., and Kacperska, A.1988. Participation of the cyanide-resis-tant pathway in respiration of winter rapeleaves as affected by plant cold acclima-tion. Physiol. Plant. 73:299–304.

Sabar, M., De Paepe, R., and de Kouchkovsky,Y. 2000. Complex I impairement, respira-tory compensations, and photosyntheticdecrease in nuclear and mitochondrial malesterile mutants of Nicotiana sylvestris.Plant Physiol. 124:1239–1249.

Salvucci, M.E. 1989. Regulation of Rubiscoactivity in vivo. Physiol. Plant. 77: 164–171.

Saradadevi, K. and Raghavendra, A.S. 1992.Dark respiration protects photosynthesisagainst photoinhibition in mesophyll pro-

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

118

toplasts of pea (Pisum sativum). PlantPhysiol. 99:1232–1237.

Saradadevi, K. and Raghavendra, A.S. 1994.Inhibition of photosynthesis by osmoticstress in pea (Pisum sativum) mesophyllprotoplasts is intensified by chilling orphotoinhibitory light; intriguing responsesof respiration. Plant Cell Environ. 17:739–746.

Saradadevi, K., Padmasree, K., and Raghavendra,A.S. 1992. Interaction between respiration,photosynthesis and photoinhibition in me-sophyll protoplasts of pea (Pisum sativum).In : Research in Photosynthesis. pp. 725–728, Vol. 3, Murata, N., Ed., Kluwer Aca-demic, Dordrecht.

Schacter, B. and Bassham, J.A. 1972 AntimycinA sensitive stimulation of rate-limitingsteps of photosynthesis in isolated spinachchloroplasts. Plant Physiol. 49:411–416.

Scheibe, R. 1990. Light/dark modulation: regu-lation of chloroplast metabolism in a newlight. Bot. Acta 103:327–334.

Scheibe, R. 1991. Redox-modulation of chloro-plast enzymes. A common principle forindividual control. Plant Physiol. 96:1–3.

Scheller, H.V. 1996 In vitro cyclic electron trans-port in barley thylakoids follows two in-dependent pathways. Plant Physiol.110:187–194.

Sharkey, T.D. 1998. Photosynthetic carbon re-duction. In : Photosynthesis: A Comprehen-sive Treatise. pp.111–122, Raghavendra,A.S., Ed., Cambridge University Press,Cambridge.

Shimazaki, K., Terada, J., Tanaka, K. and Kondo,N. 1989. Calvin-Benson cycle enzymes inguard-cell protoplasts from Vicia faba L.Implications for the greater utilization ofphosphoglycerate/dihydroxyacetone phos-phate shuttle between chloroplasts and thecytosol. Plant Physiol. 90:1057–1064.

Shyam, R., Raghavendra, A.S., and Sane,P.V. 1993. Role of dark respiration in

photoinhibition of photosynthesis and it’sreactivation in the cyanobacterium’Anacystisnidulans. Physiol.Plant. 88:446–452.

Siedow, J.N. and Day, D.A. 2000. Respirationand photorespiration. In : Biochemistry andMolecular Biology of Plants. pp. 676–728,Buchanan, B., Gruissem, W. and Jones,R., Eds., American Society of Plant Physi-ologists, Maryland.

Siedow, J.N. and Umbach, A.L. 1995. Plantmitochondrial electron transfer and mo-lecular biology. Plant Cell 7:821–831.

Siedow, J.N. and Umbach, A.L. 2000. The mi-tochondrial cyanide-resistant oxidase:structural conservation amid regulatorydiversity. Biochim. Biophys. Acta 1459:432–439.

Singh, K.K., Chen, C., and Gibbs, M. 1992.Characterization of an electron transportpathway associated with glucose and fruc-tose respiration in the intact chloroplastsof Chlamydomonas reinhardtii and spin-ach. Plant Physiol. 100:327–333.

Singh, K.K., Shyam, R., and Sane, P.V. 1996.Reactivation of photosynthesis in thephotoinhibited green alga Chlamydomo-nas reinhardtii: Role of dark respirationand light. Photosyn. Res. 49:11–20.

Stokes, D., Walker, D.A., Grof, C.P.L., andSeaton, G.G.R. 1990. Light enhanced darkrespiration. In: Perspectives in Biochemi-cal and Genetic Regulation of Photosyn-thesis. pp. 319–338, Zelitch, I., Ed., AlanR Liss, New York.

Vani, T., Reddy, M.M., and Raghavendra, A.S.1990. Beneficial interaction between pho-tosynthesis and respiration in mesophyllprotoplasts of pea during short-dark cycles.Physiol. Plant. 80:467–471.

Vanlerberghe, G.C. and McIntosh, L. 1992. Lowgrowth temperature increases alternative path-way capacity and alternative oxidase proteinin tobacco. Plant Physiol. 100:115–119.

Vanlerberghe, G.C. and McIntosh, L. 1997.Alternative oxidase: from gene to func-

Cri

tical

Rev

iew

s in

Bio

chem

istr

y an

d M

olec

ular

Bio

logy

Dow

nloa

ded

from

info

rmah

ealth

care

.com

by

110.

234.

118.

27 o

n 05

/11/

11Fo

r pe

rson

al u

se o

nly.

119

tion. Annu. Rev. Plant Physiol. Plant Mol.Biol. 48:703–734.

Vanlerberghe, G.C., Vanlerberghe, A.E., andMcIntosh, L. 1994. Molecular genetic alter-ation of plant respiration. Silencing andoverexpression of alternative oxidase intransgenic tobacco. Plant Physiol. 106:1503–1510.

Villar, R., Held, A.A., and Merino, J. 1995.Dark leaf respiration in light and darknessof an evergreen and a deciduous plantspecies. Plant Physiol. 107:421–427.

Vivekanandan, M. and Saralabai, V.C. 1997.The use of transgenic plants to manipulatephotosynthetic processes and crop yield.In : Handbook of Photosynthesis. pp. 661–669, Pessarakli, M., Ed., Marcel Dekker,New York.

Walker, D.A. 1988. The Use of Oxygen Elec-trode and Fluorescence Probes in Simple

Measurements of Photosynthesis. 2nd ed.,Oxygraphic Ltd., Sheffield, UK.

Weger, H.G. and Turpin, D.H. 1989. Mitochon-drial respiration can support NO3

– and NO2–

reduction during photosynthesis. Interac-tions between photosynthesis, respirationand N-assimilation in the N-limited greenalga Selenastrum minutum. Plant Physiol.89:409–415.

Weger, H.G., Birch, D.G., Elrifi, I.R., andTurpin, D.H. 1988. Ammonium assimila-tion requires mitochondrial respiration inthe light. A study with the green algaSelenastrum minutum. Plant Physiol.86:688–692.

Xue, X., Gauthier, D.A., Turpin, D.H., andWeger, H.G. 1996. Interactions betweenphotosynthesis and respiration in the greenalga Chlamydomonas reinhardtii. Charac-terization of light enhanced dark respira-tion. Plant Physiol. 112:1005–1014.

Cri

tical

Rev

iew

s in

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info

rmah

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care

.com

by

110.

234.

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27 o

n 05

/11/

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