how e. coli find their middle
DESCRIPTION
How E. Coli find their middle. Xianfeng Song Sima Setayeshgar. Outline. Introduction to E. coli Two systems regulate division site placement Nucleoid occlusion Min proteins Experiments: Qualitatively understand Min Proteins in E. coli Model: How and why Min Proteins work in E. coli ?. - PowerPoint PPT PresentationTRANSCRIPT
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How E. Coli find their middle
Xianfeng Song
Sima Setayeshgar
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Outline
Introduction to E. coli Two systems regulate division site placement
Nucleoid occlusion Min proteins
Experiments: Qualitatively understand Min Proteins in E. coli
Model: How and why Min Proteins work in E. coli?
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About E. coli
E. coli is a bacterium commonly found in the intestinal tracts of most vertebrates.
Studied intensively by geneticists because of its small genome size, normal lack of pathogenicity, and ease of growth in the laboratory.
Size: about 0.5 microns in diameter and 1.5 microns long
From: cwx.prenhall.com
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E. coli life cycle
Typical lifecycle
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E. coli cell division
Division accuracy: .50 +/- .02 Placement of FtsZ ring: .50 +/- .01
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Two systems regulate division site placement Nucleoid occlusion
Min proteins Including MinC, MinD, and MinE
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Min proteins (Experiments) MinC
Inhibits FtsZ ring formation
MinD MinD:ATP recruits MinC
to membrane MinE
Binds to MinD:ATP in membrane and induces ATP hydrolysis
Black: MinC Red: MinD Blue: MinE
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Min proteins (Experiments)
Without Min proteins, get minicelling phenotype (Min-)
If MinC is over-expressed, get filamentous growth (Sep-)
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Min proteins oscillate from pole to pole
Hal
e et
al.
(200
1)
MinD-GFPMinD-GFP
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MinE ring caps MinD polar region
MinE ring is membrane bound.
Ring appears near cell center and moves toward pole.
Hal
e et
al .
(200
1)
MinE-GFPMinE-GFP
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Wavelength of oscillations is ~10 microns.
Ha l
e et
al .
(200
1)
MinE-GFPMinE-GFP
Filamentous cell has “zebra stripe” pattern of oscillations
MinD-GFPMinD-GFP
Raskin and
de Boer (1999)
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Phenomenology of Min oscillations MinD polar regions grow as end caps. MinE ring caps MinD polar region. Oscillation frequency:
[MinE] frequency , [MinD] frequency .
Filamentous cell has “zebra stripe” pattern of oscillations.
In vivo oscillations require MinD and MinE but not MinC.
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How and Why Min-proteins oscillate (Modeling effort) Howard et al. (2001)
Very simple 1D model MinE is recruited by cytoplasmic MinD to membrane MinD polar region fails to reform at poles. (not agree with experi
ment) Huang and Wingreen (2003)
MinE is recruited by membrane-bound MinD-ATP MinD aggregation on the membrane follow a one-step process
Kruse et al. (2005) Consider protein diffusion on the membrane. MinD aggregation on the membrane follow a two-step process: fir
st attach to membrane, then self-assembles into filament Meinhardt and de Boer (2001)
Requires new protein synthesis.
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Model from Huang and Wingreen (2003)
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Equations for model from Huang and Wingreen (2003)
membranein ATP:MinD:MinE
membranein ATP:MinD
cytoplasmin MinE
cytoplasmin MinD
de
d
E
D
dedeEdEde
dt
d
EdEdedeEEE
dt
d 2D
ATPDdeddDDEdEd
dt
d:)]([
s
ss
ss
ED
deE
dDD
2
3
3
m 5.2
1 8.0;
m 16.0
m001.0;
m025.0
DD
ATPDdeddDD
ATPADP
ADPDATPDD
ATPD
dt
d
:
::
2:
)]([
D
ATPADP
ADPDdedeADPDD
ADPD
dt
d
:
:2: D
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Result: MinD/E movie
MinE
MinDMinD
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Result: MinD end caps and MinE ring
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Mechanism for growth of MinD polar regions (Huang and Wingreen, 2003) MinD ejected from old end cap
diffuses in cytoplasm.
Slow nucleotide exchange implies uniform reappearance of MinD:ATP.
Capture of MinD:ATP by old end cap leads to maximum of cytoplasmic MinD:ATP at opposite pole.
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Result: Frequency of oscillations ~ [MinE]/[MinD]
No oscillations for [MinE] too high, or for [MinD] too low.
Minimum oscillation period 25s.
4 micron cell 4 micron cell
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Result: “Zebra stripe” oscillations in long cells
Stripes form with wavelength of ~10 microns
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Oscillation makes E. Coli divide accurately The oscillation make MinD has smaller avera
ge concentration at the middle and higher concentration at the ends.
MinC can only be bounded to memberane if it is recruited by MinD-ATP
MinC can Inhibits FtsZ ring formation, thus inhibits the division
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Linear stability analysis around homogeneous solution
tikziii etx 0),(
Red curve corresponds to a normal cell
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Predictions of model (Huang and Wingreen, 2003)
Delay in MinD:ATP recovery is essential. (Verified by Joe Lutkenhaus – nucleotide exchange rate is slow, ~2 / s.)
Rate of hydrolysis of MinD:ATP by MinE sets oscillation frequency.
Diffusion length of MinD before rebinding to membrane sets oscillation wavelength.
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Recent development Helical morphology of MinD polymers.
Some recent modeling effort (mainly focus on cleaning some details, no breakthrough) the fluctuation effects on the min proteins (Howard, et al, 2003), The new model considering the membrane diffusion and more reactions
(Kruse, et al, 2005) min-protein oscillations in round bacteria (Huang and Wingreen, 2004)
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Open questions
MinE ring moving in reverse.
The helical structure of MinD polymers
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Conclusions
Although bacteria such as E. coli is a very simple creature, it is also very complicated system.
To understand this system, physicist can help a lot.
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Thanks to Doc. Wingreen kindly let us to use his slides.
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Thank you
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Evidence from in vitro studies
A. Phospholipid vesicles
B. MinD:ATP binds to vesicles and deforms them into tubes
C. MinD:ATP polymerizes on vesicles
D. Diffraction pattern indicates
well-ordered lattice of MinD:ATP
E. MinE induces hydrolysis of MinD:ATP and disassembly of tubes
Hu
Hu et al.
et al. (2002) (2002)
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Min proteins in spherical cells:Neisseria gonorrhoeae
Wild typeWild typeMinDMinDNgNg
--
Sze
to
Sze
to e
t al
et a
l . (2
001)
. (2
001)
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Min-protein oscillations in nearly round cells
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In E. coli, Min oscillations “target” MinD to poles
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Why does E. coli need an oscillator?
In B. subtilis, minicelling is prevented by MinCD homologs, but polar regions are static.
Marston Marston et alet al. (1998). (1998)
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How E. coli find its middle Proteins are too small to see the caps’ curvature Subtilis have local proteins fixed at two ends, but
E. coli does not have