[CANCER RESEARCH 30,1075-1080,April1970]

SUMMARY

Morris hepatoma S123tc in intact or adrenalectomizedBuffalo strain rats has a very low basal level of tryptophanpyrrolase (TP) activity and a high basal level of tyrosinetransaminase (TKT) activity in comparison with normal liver.Both enzymes are inducible by hydrocortisone injections,but the induced level of TP in the hepatoma is much lessthan that of liver. Injections i.p. of histones or poly-L-lysineinduce increases in TP, but not TKT, in the hepatoma inadrenalectomized rats.

The rate of decline of induced 1? activity in the S123tchepatoma in the presence of puromycin to prevent furthersynthesis of the enzyme is identical with the rate of declinein normal liver, and consequently the low basal levels andweak inducibiity of TP in this tumor probably should beattributed to a defect in synthesis of TP, rather than toaccelerated rate of degradation of this enzyme. Injectedhistones decrease the rate of decline of TP activity in thehepatoma as well as liver. On the other hand, there is nodecline of induced TKT activity in the hepatoma during an8-hr period after induction, in contrast to normal liver, inwhich the half-life of TKT is 2.9 hr. Consequently, the highbasal level of TKT in this hepatoma probably should beattributed to a low rate of degradation rather than a highrate of synthesis. The rate of degradation of TKT in liver ofrats bearing this hepatoma also is greatly decreased incomparison with normal liver; this suggests that the tumormay produce a circulating inhibitor of the degradative system for TKT.

1Aided by Grant GM 08318-08 from the National Institute ofGeneral Medical Sciences Grant CA-10729 from the National CancerInstitute, NIH, and Grant E-241 from the American Cancer Society.Some of the equipment used in this research was obtained throughDepartmental Equipment Grant GB-4577 from the National ScienceFoundation.

2@OGto@l Research Trainee, supported by Training Grant5T01GM00404 from the National Institute of General MedicalSciences, NIH.

Received February 24, 1969; accepted October 16, 1969.

Basal levels of TP and TKT and inducibility by hydrocortisone and poly-L-lysine were studied in four other Morrishepatomas: 5l23A, 5l23C, 7793, and 7795.

INTRODUCTION

Induction of tryptophan pyrrolase activity in livers ofintact tumor-bearing rats and 14 “minimaldeviation― (19)transplantable hepatomas was studied by Dyer et aL (5) withL-tryptophan as the inducer. Some of the hepatomas wereobserved to have very low TP@ activities (e.g., 5 123tc,5123C) and to respond minimally (5 123C), or not at all(5 l23tc), to induction by tryptophan. Dyer Ct al. (5) ruledout the possibility that insufficient inducer reached thehepatoma by inducing the enzyme in hepatoma H35 (20)growing in the “tissueisolated― kidney (8) and by introducing the inducer directly into the kidney artery. No greateractivity was found in either the tumor or the host liver bythis procedure than occurred following i.p. injection of theinducer. Pitot and Morris (1 7) also showed that adequateinducer was available to the tumor. Cho and Pitot (3) andCho et a!. (4) tested the possibility that the structure of theenzyme in the hepatoma might be different from that of thehost liver, and they found no significant difference betweenthe partially purified enzymes of liver and hepatoma 7793with respect to the kinetic constants or the electrophoreticmobiities. Partially purified TP from hepatoma H35 also hada similar Km value to that of the liver enzyme.

Schimke et a!. (22, 23) have reported that the level ofactivity of an inducible enzyme is the result of the rates ofsynthesis and degradation of the enzyme. The induction ofan enzyme (e.g., TP) can occur either through stimulation ofenzyme synthesis (e.g., effect of hydrocortisone) or throughdecrease in rate of degradation (e.g., effect of tryptophan).In the light of this hypothesis, it seemed important todetermine whether the variations in basal levels and induci

3The abbreviations used are: TP, tryptophan pyrrolase(L-tryptophan:oxygen oxidoreductase, EC 1.13.1 .12); TKT, tyrosinea-ketoglutarate transaminase (L-tyrosine :2-oxoglutarateaminotransferase, EC 2.6.1.5).

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Effects of Exogenous Histones upon the Induction of TryptophanPyrrolase and Tyrosine Transaminase in Liver and Morris

1

Henry W. Strobel, Jr.,2 J. Logan Irvin, and Harold P. Morris

Department of Biochemistry, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27514 fH. W. S., .1. L. 1.1 andDepartment of Biochemistry, CollegeofMedicine, Howard University, Washington,D. C. 20001 /H. P. M./

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H. W. Strobe!, Jr. , J. L. Irvin, and H. P. Morris

M) by a modification of the method of Knox and Mehler

(1 0) and Feigelson and Greengard (@) after a 30-mm preincubation (1 1) of the homogenate at room temperature withsubstrate . Tyrosine transaminase activity was determined at370 by the method of Rose et a!. (2 1) and is expressed aspmoles p-hydroxyphenylpyruvate/min/g fresh liver.

RESULTS

Comparison of Various Hepatomas in NonadrenalectomizedRats. The data of Chart 1 show the induction by hydrocortisone and poly-L-lysine of TP and tyrosine transaminase in SMorris hepatomas in nonadrenalectomized hosts. Both TPand TKT are inducible by hydrocortisone and poly-L-lysinein hepatoma S 123tc. The enzymes differ in response in thatthe 0.9% NaCI solution-injected control level of tumor TP isreduced much below the 0.9% NaCl solution-control level innormal liver, while the NaCl solution control level of tumorTKT is elevated in comparison with normal liver. Hepatomas5123C, 5l23A, and 7795 show reduced NaCl solutioncontrol levels of TP and no significant inducibility byhydrocortisone or poly-L-lysine. Hepatoma 7793 shows onlya slightly depressed saline control level of TP in comparisonwith liver and a slight induction by hydrocortisone andpoly-L-lysine (p 0.03 and 0.02, respectively). HepatomasS 123C and S123A show large values of TKT activities in0.9% NaCl solution-injected controls in comparison withliver. In fact, the basal level TKT activities in these hepatomas are almost as high as those in hydrocortisone-inducednormal liver. TKT cannot be induced to higher levels bypoly-L-lysine in either S l23C or S l23A, nor by hydrocortisone in S l23C. Hepatoma 7795 shows an elevated 0.9%NaCl solution control level of TKT and slight inducibility byhydrocortisone and poly-L-lysine (p 0.02 and 0.03, respec

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Chart 1. Induction of TP and tyrosine transaminase of normal liverand various Morris hepatomas in nonadrenalectomized Buffalo strainrats. The rats were sacrificed 5 hr after injection of 0.5 ml 0.9% NaC1solution (0), 2 mg hydrocortisone 21-acetate in 0.5 ml 0.9% NaClsolution (Iii]), or 10 mg poly-L-lysine hydrobromide in 0.5 ml 0.9%NaCl solution (a) per rat (average weight, 200 g). TP activity is inj.imoles kynurenine/hr/g liver. TKT activity is in @@molesp-hydroxy.phenylpyruvate/min/g liver. Bars, average values for at least 6 experiments with each tumor. S.E. (T-bars) was calculated for applicationof the t test, and the statistical significancesof comparisons are citedin the text.

biity of TP and TKT in Morris hepatomas are attributableto defects in synthesis or degradation of these enzymes incomparison with normal liver. This paper reports a study ofthis type with I hepatoma (5 123tc).

Caffery et a!. (2) found that hepatic TP and TKT areinduced in adrenalectomized mice by i.p. injection ofhistones or poly-L-lysine, but paradoxically the injection ofhistones 30 to 60 mm before hydrocortisone inhibits theinduction of TP and TKT, which otherwise would resultfrom either injection given singly. In the present paper, theeffects of injected histones and poly-L-lysine upon inductionof TP and TKT of several hepatomas are compared witheffects upon induction of these enzymes in liver of thetumor-bearing rats, as an additional means of evaluating thevariations in the inductive mechanisms of the hepatomas.

MATERIALS AND METHODS

The hepatoma lines used in these studies have beendescribed previously (13, 15, 16). All tumors had beeninduced by N-2-fluorenylphthalamic acid. The 5 l23tc linehad been grown in tissue culture at the McArdle Laboratoryfor Cancer Research for several months and then returned tothe rat. The tumors were received in the North Carolinalaboratory as intramuscular implants in the hind legs ofBuffalo strain rats. The tumor lines were maintained byinoculation of a 50% suspension of hepatoma cells in 0.9%NaCl solution into the axillary regions of 80 to 100 g maleBuffalo strain rats obtained from Simonsen Laboratories,Gilroy, Calif. The tumors grew well in the axillary region,could be checked conveniently for growth by palpation, andcould be excised easily at sacrifice.

When required, rats were bilaterally adrenalectomized 3 to5 days before sacrifice by use of the technique originallydescribed by Grollman (7). The adrenalectomized rats weremaintained on Purina chow and 0.9% NaCl solution-5.0%giucose in place of drinking water. All injections wereadministered i.p. dissolved or suspended in 0.9% NaC1 solution. PoIy-L-lysine hydrobromide and hydrocortisone 21-acetate were obtained from Sigma Chemical Company, SaintLouis, Mo.; puromycin dihydrochioride was obtained fromNutritional Biochemicals Corporation , Cleveland, Ohio.Unfractionated calf thymus histones were prepared by themethod of Bellair and Mauritzen (1) and were checked forpurity by amino acid analyses [by the method of Spackmanet a!. (24)1 and polyacrylamide gel electrophoresis ( I 2) inthe presence of 6 M urea.

Rats were sacrificed 5 hr after injection, and the livers wereimmediately removed, blotted dry, weighed, and frozen.Tumors were then excised, freed of connective and necrotictissue, weighed, and frozen. Homogenates were prepared bydiluting the liver or tumor with 7 volumes alkaline KCI (0.14M KC1-2.5 mM NaOH) and homogenizing (final pH, 7.0) in a

Potter-Elvehjem glass homogenizer with a Teflon pestle. Thehomogenates were filtered through several thicknesses ofcheesecloth. TP activity, expressed as pmoles kynurenine/hr/g fresh liver, was assayed at 370 with optimalconcentrations of hematin (0.5 pM) and ascorbic acid (0.01

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Enzyme Induction in Liver and Hepatomas

induction of TP activity by hydrocortisone, in host liver by40% and tumor by 60%, in comparison with data for thecorresponding group injected with hydrocortisone alone.

Adrenalectomy does not change the 0.9% NaCl solutioncontrol level of host liver tyrosine transaminase, but it doesdecrease the hydrocortisone-induced activity and it doeseliminate the poly-L-lysine induction seen in nonadrenalectomized rats. The 0.9% NaC1 solution control level of tumorTKT is lower in adrenalectomized rats than in nonadrenalectomized animals, while the hydrocortisone-induced level isnot significantly different in the 2 groups. However, therelative induction of TKT is approximately 3-fold greater inthe S123tc tumors of adrenalectomized rats than in tumorsof nonadrenalectomized rats. Induction by poly-L-lysine oftumor TKT is not significant either in nonadrenalectomizedor adrenalectomized rats, in comparison with the corresponding 0.9% NaCl solution-injected controls. Histones injectedalone into tumor-bearing adrenalectomized rats have noeffect on TKT activity in the tumor and seem to cause somedecrease of activity in host liver. A single injection ofhistones 30 mm before hydrocortisone inhibits the hydrocortisone induction of TKT in the tumor by 30%, while the

inhibition in host liver (1 1%) is not statistically significant.Rates of Decline of Induced TP and TKT Activities in

Liver and Hepatoma 5 123tc and Effects of Injected

lively), while 7793 shows a reduced 0.9% NaCl solutioncontrol level and a slight inducibility by poly-L-lysine, butnot by hydrocortisone (p 0.01 and 0.1 , respectively).

Detailed Study of Hepatoma 5 123tc. Hepatoma S123tc waschosen for a more detailed study, since both enzymes areinducible in this tumor and since I enzyme (TP) has a 0.9%NaC1 solution-injected control level below that of normalliver and the other enzyme (TKT) has a 0.9% NaCI solutioncontrol level elevated above that of normal liver.

The data of Chart 2 emphasize the involvement of adrenalfunction in enzyme induction. A significant decline isevident in the 0.9% NaCl solution control, hydrocortisoneinduced, and poly-L-lysine-induced activities of host liver TPafter adrenalectomy. In fact, the statistically significantinduction by poly-L-lysine of host liver TP in nonadrenalectomized animals is eliminated by adrenalectomy. Thisdependence on adrenal function is seen also in the tumor,where a statistically significant low level of TP activity (0.9%NaCI solution controls) vanished upon adrenalectomy.Induction of tumor TP by hydrocortisone is not significantlychanged by adrenalectomy, while induction by poly-L-lysineis increased. Histones injected alone also seem to increasetumor 1? activity in adrenalectomized rats, but injectedhistones have no effect on host liver TP activity. Histonesinjected 30 mm before hydrocortisone significantly inhibit

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Chart 2. Induction by hydrocortisone, poly-L-lysine,and histones of TP and TKT in host liverand tumor of rats bearing hepatoma 5 l23tc, and the effect of injected histones uponhydrocortisone induction of these enzymes in adrenalectomized and nonadrenalectomizedBuffalo strain rats. Unfractionated calf thymus histones (15 mg/ 100 g body weight) wereinjected (in 0.5 ml 0.9% NaC1 solution) into the histone-treated and into the histone-hydrocortisone acetate-treated groups. In the latter group, histones were injected 30 mm prior tohydrocortisone acetate (HO (5 mg/rat). The rats had average body weights of 180 g at the timeof these experiments. Animals were sacrificed for enzyme assays S hr after injection of 0.9%NaCl solution, hydrocortisone, poly-L-lysine, or histones, as in Chart 1. Enzyme units are thesame as those in Chart 1, but note that the TP activity scale for the tumor is 0.1 that of hostliver. A,@ , host liver of nonadrenalectomized rats; B,@ ,host liver of adrenalectomized rats; C,l:@B, tumor of nonadrenalectomized rats; D,@ , tumor of adrenalectomized rats. T-bars, S.E. for 7to 10 rats/group in the nonadrenalectomized series and 4 to 9 rats/group in the adrenalectomized series.

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APRIL 1970 1077

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H. W. Strobe!,Jr.,J.L.Irvin,and H. P.Morris

Histones. Experiments similar in design to those of Schimkeet a!. (22, 23) were performed to determine the rates ofdecline of induced TP and TKT activities in normal liver ofnontumor-bearing adrenalectomized rats and in host liver andtumor of adrenalectomized rats bearing the S123tc hepatoma. The results are recorded in Charts 3 and 4. In theseexperiments, the activities of TP and TKT were induced torelatively high levels by 2 i.p. injections (4 hr apart) ofhydrocortisone acetate (5 mg) and L-tryptophan (100 mg)into each rat in the various groups. Five hr after the lastinjection, designated as 0 time, experimental rats received ani.p. injection of histones, while control rats received 0.9%NaC1 solution. Puromycin was injected into all animals atindicated points to prevent synthesis of TP and TKT duringthe period of measurement of the decline in enzyme activity.The data in Chart 3 indicate that host liver TP activity,induced 10-fold over basal activity (Chart 2) by injection ofhydrocortisone and tryptophan, declines at a rate (t½= 3.5hr) 0.7 hr slower than induced TP activity in normal liver(t½ 2.8 hr). A single injection of histones increases the

half-life of host liver TP by 13 hr, which is comparable withthe 1-hr extension of normal liver TP half-life by injectedhistones. Tumor TP activity, induced approximately 10-foldover basal tumor TP activity by the prior i.p. injections ofhydrocortisone and tryptophan, declines at a rate (t½= 2.7hr) almost identical with that of induced normal liver TPactivity (t½ 2.8 hr). A single injection of histones extendsthe half-life of tumor TP to 3.6 hr, identical with thehalf-life extension by histones of TP activity in inducednormal liver.

The data of Chart 4 show that host liver tyrosine transaminase activity, induced 4-fold over basal host liver TKTactivity by injection of hydrocortisone and tryptophan,declines with a half-life almost 10 hr longer than inducedTKT activity in normal liver (t½ = 2.9 hr). A singleinjection of histones eliminates the decline in host liver TKTactivity over the time course for this experiment. TumorTKT activity is induced approximately 4-fold over basaltumor TKT activity in adrenalectomized rats and shows nosignificant decline during the course of this experiment. Asingle injection of histones has no effect on the rate ofchange of tumor TKT activity as shown in Chart 4.

DISCUSSION

The data presented in this paper (Chart 1) for the activitiesof tryptophan pyrrolase and tyrosine transaminase for aspectrum of Morris hepatomas in intact rats agree well withprevious work (4, 5 , 18). The basal levels of these enzymesand the response of these enzymes to induction by hydrocortisone vary greatly from hepatoma to hepatoma. In somehepatomas (S I 23A and 7795) basal level TP is reducedbelow that of normal liver and shows no induction byhydrocortisone, while basal level TKT activity is elevatedabove that of normal liver and shows a small degree of

Other hepatomas have a wideof elevation or reduction ofor absence of induction by

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Chart 3. The effect of i.p. injection of unfractionated calf thymushistones on the rate of decline of induced TP activity of normal liverof nontumor-bearing adrenalectomized rats and host liver and tumorof adrenalectomized rats bearing the 5123tc hepatoma. High levels ofTP activity were induced in each rat by 2 i.p. injections (4 hr apart)of hydrocortisone acetate (5 mg) and L-tryptophan (100 mg) per rat(average body weight, 190 g). Five hr after the last injection,designated as 0 time, experimental rats received 1 injection ofhistones (15 mg/100 g body weight) while control rats received 0.9%NaG solution injection. All animals of control and experimentalgroups received puromycin hydrochloride (6 mg/rat by i.p. injectionat points designated by P and arrows). Rats from control andexperimental groups were sacrificed at designated time periods for TPassay, and averages of the data for 6 rats/group were recorded. I-bars,S.E. ., U, A, control (0.9% NaCl solution-injected) rats; a, o, @,histone-injected rats. A, t@,liver of nontumor-bearing rats; ., o, tumor;and ., o, livers of tumor-bearing rats.

Chart 4. The effect of i.p. injection of unfractionated calf thymushistones on the rate of decline of induced TKT activity of normalliver of nontumor-bearing adrenalectomized rats and host liver andtumor of adrenalectomized rats bearing the 5 123tc hepatoma. Theanimals in this experiment were identical with those described in thelegend to Chart 3, and the symbols have identical significance.

A more detailed study of these enzymes in a singlehepatoma in comparison with normal and host liver seemednecessary. In the course of the study of hepatoma 5123tc(Chart 2), it was observed that the induction by poly-L

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induction by hydrocortisone.variety of the combinationsbasal activity and inductionhydrocortisone.

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Enzyme Induction in Liver and Hepatomas

lysine of hepatic TP and TKT which was found in nonadrenalectomized tumor-bearing rats was not present inadrenalectomized tumor-bearing rats. The elimination ofinduction by poly-L-lysine through adrenalectomy suggeststhat poly-L-lysine induction of TP and TKT in liver of theintact rat is an adrenally mediated phenomenon, possibly astress reaction. In contrast to these results for the rat,Caffery et a!. (2) have found significant “induction―ofhepatic TP and especially TKT by i.p. injection of poly-Llysine and histones into adrenalectomized mice, but thisinduction was attributed to decreased rate of degradation ofthese enzymes rather than to stimulation of synthesis.

The induction of TP in hepatoma S l23tc by poly-L-lysineand histones injected into adrenalectomized rats cannot beinterpreted as an adrenally mediated increase in synthesis.Schimke et a!. (22, 23) have proposed that induction ofenzyme activity is the result of factors affecting the rate ofenzyme degradation and enzyme synthesis. Thus, an increasein enzyme activity could be the result of an increase in therate of synthesis, a decrease in the rate of degradation, orboth. The data of Chart 3 show that injected histonesdecrease the rate of degradation of TP in the hepatoma, andconsequently the induction of TP in hepatoma S123tc bypoly-L-lysine and histones may be explained as the result ofa decrease in the rate of degradation of TP. However, somestimulation of synthesis of TP is not excluded by theseexperiments. The disappearance of hepatoma 1? activity in0.9% NaCl solution-injected controls after adrenalectomy andthe low levels of hydrocortisone-induced TP activity inhepatoma 5 l23tc (Chart 2) can be interpreted as a markeddecrease in the rate of synthesis of this enzyme, since therate of decline of TP activity in the hepatoma is not

significantly different from that of normal liver (Chart 3).This interpretation is consistent with the hypothesis of Choet a!. that the hepatoma may have lost the capacity tomaintain a stable RNA template for TP synthesis (4).

The observations from Chart 3 that induced tumor TPactivity declines at the same rate as induced TP activity innormal liver and that a single injection of histones extendsthe half-life of tumor TP and normal liver TP to the sameextent suggest that the degradative mechanism is normal andthat the synthetic mechanism for tumor TP may be the siteof the defect in the regulation of TP in this hepatoma. Onthe other hand, the observations (Charts 2 and 4) that tumortyrosine transaminase activity in the induced state does notdecline significantly and that tumor TKT is inducible suggestthat the degradative mechanism for tumor TKT may be thesite of the defect in the regulation of TKT in this hepatoma.Apparently, the degradative mechanism for TKT is stronglyinhibited or absent in this tumor, since no decline in activitywith time is observed under conditions in which furthersynthesis is blocked by puromycin. Injected histones have noeffect on the absence of decline in tumor TKT activity . Bothdecline in activity of TKT with time and extension ofhalf-life of this enzyme by injected histones are observed forhost liver in the same animals. However, the rate of declineof induced TKT activity in host liver (Chart 4) is muchslower than in liver of nontumor-bearing adrenalectomizedrats (t½ 12 .5 hr and 2 .9 hr, respectively). This observation

suggests the possibility that hepatoma S123tc may produce adiffusible compound which decreases the rate of degradationof TKT, not only in the tumor itself, but also in the liversof rats bearing this hepatoma. Neither the nature of this“stabilizer―of TKT nor its mode of action is known. Thiseffect of the tumor may be an indirect one, and it ispossible that the stabilizer may be produced as a result of anaction of the tumor upon another tissue of the host or as aresult of the generalized cachexia resulting from the tumor.If synthesis of TKT continued in tumor and host liver underconditions of a low rate of degradation, a continuously risingactivity of TKT in liver and tumor might be expected in theabsence of puromycin to block further synthesis. However,the TKT activities reach plateau levels after an injection ofhydrocortisone and do not increase continuously. Consequently, there must also be a mechanism to stop furthersynthesis of TKT. Degradation of the mRNA's for TKTsynthesis is 1 possible mechanism for this limitation on therise in TKT activity. The observations on rates of decline ofinduced TP and TKT activities in hepatoma 5 123tc are notnecessarily applicable to other Morris hepatomas.

The mechanism by which the injected histones produce adecrease in the rate of decline of TP and TKT activities isnot known. However, Kenney (9) has concluded from hisexperiments that a continuously synthesized polypeptide(probably with short half-life) participates in the degradativephase of TKT turnover. It seems possible that injectedhistones may inhibit the synthesis of the polypeptide orpolypeptides which are involved in the degradation or mactivation of TP or TKT, and this inhibition of degradationwould account for the effect of injected histones in prolonging the half-lives of TP and TKT in liver (Charts 3 and4). The production of the degradative polypeptide for TKTmay be inhibited already in the hepatoma and thereforewould be unaffected by injected histones.

The repression of hydrocortisone induction of host liverand tumor TP and TKT by histones injected 30 mm beforehydrocortisone (Chart 2) may be interpreted as a modification of the rate of synthesis of TP and TKT, according toSchimke's hypothesis. Histones may inhibit the productionof the RNA templates for TP and TKT. This view isconsistent with the work of Moore and Irvin (14), whoshowed inhibition of RNA synthesis by injected histones inthe livers of adrenalectomized mice. Although this interpretation of the action of injected histones is attractive, it is aworking hypothesis, and there are other possible mechanismsof action, such as modification of membrane permeabilitiesor stimulation or repression of the secretion of hormonesother than adrenal corticoids. However, experiments withmice in this laboratory (J. M. Caffery and J. L. Irvin,unpublished results) with doubly labeled histones haveindicated that i.p. injected histones do appear in nuclei ofliver cells in these animals. The uptake into cell nuclei is asmall percentage of the injected quantity, but the smallamount does not necessarily rule out this effect, since asimilarly small quantity (1 5 to 20 pg) of actinomycin D inhibitsRNA synthesis in mice.

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H. W. Strobe!, Jr., J. L. Irvin, and H. P. Morris

13. Miyaji, H., Morris, H. P., and Wagner,B. P. HistologicalStudy ofSome Primary and Transplantable Hepatic Tumors in the Rat. In:H. Busch (ed.), Methods in Cancer Research, Vol. 4, pp.153 —178. New York: Academic Press, Inc., 1968.

14. Moore, P. A., and Irvin, J. L. Inhibition of Liver RNA Synthesisby Injected Histones. Federation Proc, 25: 645, 1966.

15. Morris, H. P. Some Growth, Morphological and BiochemicalCharacteristics of Hepatoma 5123 and Other New TransplantableHepatomas. hog. Exptl. Tumor Res., 3: 370—411,1963.

16. Morris, H. P., and Wagner, B. P. Induction and Transplantationof Rat Hepatomas with Different Growth Rate (including “Minimal Deviation― Hepatomas). In: H. Busch (ed.), Methods inCancer Research, Vol. 4, pp. 125—152.New York: AcademicPress, Inc., 1968;

17. Pitot, H. C., and Morris, H. P. Metabolic Adaptations in RatHepatomas. II. Tryptophan Pyrrolase and Tyrosine a-Ketoglutarate Transaminase. Cancer Res., 21: 1009—1014,1961.

18. Pitot, H. C., Peraino, C., Bottomley, R H., and Morris, H. P. TheComparative Enzymology and Cell Origin of Rat Hepatomas. III.Some Enzymes of Amino Acid Metabolism. Cancer Res., 23:135—142,1963.

19. Potter, V. R. Transplantable Animal Cancer, the PrimaryStandard.CancerRes.,21: 1331—1333,1961.

20. Reuber, M. D. A Transplantable Bile-secreting HepatocellularCarcinoma in the Rat. J. Natl. Cancer Inst., 26: 891—897, 1961.

21. Rosen, F., Harding, H. R., Milholland, R J., and Nichol, C. A.Glucocorticoids and Transaminase Activity. VI. Comparison ofthe Adaptive Increases of Alanine and Tyrosine-x-KetoglutarateTransaminase. J. Biol. Chem., 238: 3725—3729, 1963.

22. Schimke, R. T., Sweeney, E. W., and Berlin, C. M. An Analysisof the Kinetics of Rat Liver Tryptophan Pyrrolase induction:The Significance of Both Enzyme Synthesis and Degradation.Biochem.Biophys.Res.Commun.,15:214—219,1964.

23. Schimke, R. T., Sweeney, E. W., and Berlin, C. M. The Roles ofSynthesis and Degradation in the Control of Rat Liver Tryptophan Pyrrolase. J. Biol. Chem., 240: 322—331, 1965.

24. Spackman, D. H., Stein, W. H., and Moore, S. AutomaticRecording Apparatus for Use in the Chromatography of AminoAcids Anal. Chem., 30: 1190—1206, 1958.

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2. Caffrey, J. M., Whichard, L., and Irvin, J. L. Induction ofHepatic Tryptophan Pyrrolase and Tyrosine Transaminase byHistones and other Polypeptides. Biochim. Biophys. Acta, 157:616—626,1968.

3. Cho, Y. S., and Pitot, H. C. Substrate-Hormone Relationshipsduring Tryptophan Pyrrolase Induction in Liver and Hepatomas.Proc. Am. Assoc. Cancer Res., 4: 11, 1963.

4. Cho, Y. S., Pitot, H. C., and Morris, H. P. Metabolic Adaptationsin Rat Hepatomas. VI. Substrate-Hormone Relationships inTryptophan Pyrrolase Induction. Cancer Res., 24: 52—58, 1964.

5. Dyer, H. M., Gullino, P. M., and Morris, H. P. TryptophanPyrrolase Activity in Transplanted “MinimalDeviation― Hepatomas. Cancer Res., 24: 97—104, 1964.

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1970;30:1075-1080. Cancer Res   Henry W. Strobel, Jr., J. Logan Irvin and Harold P. Morris  Morris HepatomasTryptophan Pyrrolase and Tyrosine Transaminase in Liver and Effects of Exogenous Histones upon the Induction of

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