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Biomedical subjects

B Peterkofsky

Publications and source records attributed to B Peterkofsky.

At least 37 records · Page 2Linked to original sources
PubMed1988

Salt stimulation of serum insulin-like growth factor binding protein activity.

Insulin-like growth factors (IGF)-I and -II are bound to carrier or binding proteins in serum. There are at least two classes of binding protein: a high molecular weight complex and a low molecular weight species that is relatively unsaturated. Total binding capacity in serum generally is determined by incubating [125I]IGF with protein that has been stripped of IGF by acid gel filtration. We found that addition of NaCl to the assay increased binding to stripped guinea pig binding protein to about two to four times the level measured in the absence of salt. Stimulation by NaCl was optimal between concentrations of 0.6 and 1.4 M and also was observed when fetal calf or human sera were used as sources of stripped binding protein or when IGF-II was the ligand. Using chloride salts, the order of activity with respect to cations was Na+ greater than K+ greater than Li+. Na2HPO4 at 0.6 M was as stimulatory as 1.2 M NaCl but 0.6 M Na2SO4 was less effective. NH4HCO3 was as effective as NaCl at 0.6 M. Scatchard plots of data from competitive dilution experiments with [125I]IGF-I and unlabeled IGF-I showed that binding was heterogeneous in the absence of 0.6 M NaCl but linear in its presence. NaCl did not stimulate binding when whole serum was used, but after gel filtration of serum on Sephacryl 200 at pH 8, which does not dissociate IGFs from binding protein, binding to individual fractions was stimulated three- to fourfold by NaCl. Fractions stimulated included those containing the large complex or the unsaturated binding protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

The effectiveness of inhibitors of soluble prolyl hydroxylase against the enzyme in the cisternae of isolated bone microsomes.

Inhibitors of purified, soluble prolyl hydroxylase (K. Majamaa et al. (1984) Eur. J. Biochem. 138, 239-245; K. Majamaa et al. (1986) J. Biol. Chem. 261, 7819-7823) were tested against isolated chick embryo bone microsomes containing intracisternal prolyl hydroxylase and its radiolabeled, unhydroxylated procollagen substrate. Two groups of inhibitors were used which consisted of pyridine-2-carboxylate and 1,2-dihydroxybenzene (catechol) derivatives. The 2,4- and 2,5-pyridine dicarboxylic acids, which are potent inhibitors of the soluble enzyme (Ki values 2 and 0.8 microM, respectively), were effective in the same concentration range against intracisternal prolyl hydroxylase, although their relative affinities were reversed. Inhibition by pyridine-2,4-dicarboxylate in the microsomal system was reversed by increasing the concentration of 2-oxoglutarate. Pyridine-2,4-dicarboxylic acid did not inhibit the uptake of 2-[14C]oxoglutarate into microsomes, so it appears likely that the inhibitor must traverse the microsomal membrane and act directly at the enzyme level. Pyridine-2-carboxylic acid was ineffective in the microsomal system at 1 mM whereas it is a relatively potent inhibitor of the soluble enzyme with a Ki of 25 microM. This finding suggests that the second carboxyl group of the pyridine carboxylate derivatives may be required for their transport into the microsomal lumen. In the soluble system, 3,4-dihydroxybenzoic acid and 1,2-dihydroxybenzene had been found to be competitive inhibitors with relatively low Ki values of 5 and 25 microM, respectively. In the microsomal system, half-maximal inhibition was obtained at approximately 50-100 microM and inhibition was not reversed by increasing the concentrations of either 2-oxoglutarate or ascorbate, alone or together. These results imply that in situ these compounds do not inhibit prolyl hydroxylase directly. Thus, the microsomal system can assess the accessibility of the intracisternal enzyme to potential inhibitors and offers an insight into the in cellulo potential of such compounds.

Animals↗
PubMed1987

Iron-dependent uptake of ascorbate into isolated microsomes.

A preliminary study (J.M. Mata, R. Assad, and B. Peterkofsky (1981) Arch. Biochem. Biophys. 206, 93-104) suggested that chick embryo limb bone microsomes took up and concentrated [14C]ascorbate in the presence of cofactors for prolyl hydroxylase. In the present study, we found that the apparent Km for ascorbate in the hydroxylation of intracisternal unhydroxylated procollagen by endogenous prolyl hydroxylase was approximately an order of magnitude less than the value obtained when enzyme solubilized from microsomes was used with an exogenous substrate. These results are compatible with a concentrative uptake of ascorbate into microsomes. The uptake of [14C]ascorbate into microsomes was confirmed and it required only iron, in either the ferrous or ferric form, and was time and temperature dependent, proportional to microsome concentration, and substrate saturable at 2-3 mM ascorbate. Iron-dependent ascorbate uptake also was observed with L-929 cell microsomes. [14C]Ascorbate seemed to be taken up without prior oxidation, since only unlabeled ascorbate, and not dehydroascorbate, competed for uptake into limb bone microsomes. A functional requirement for Fe2+ in ascorbate transport was demonstrated using the intracisternal proline hydroxylating system. L-929 cell microsomes were preincubated with ascorbate with or without the metal and then external ascorbate was oxidized to inactive dehydroascorbate using ascorbic acid oxidase, which cannot penetrate the microsomal membrane. Samples which did not receive iron during the preincubation received it, along with other requirements for prolyl hydroxylase, in a final incubation to measure hydroxylation. Significant hydroxylation was obtained only in samples incubated with iron prior to oxidase treatment, consistent with the conclusion that an iron-dependent process was required to translocate ascorbate and protect it from the oxidase.

Animals↗
PubMed1986

Suppression of synthesis of pro-alpha 1(I) and production of altered pro-alpha 2(I) procollagen subunits in 4-nitroquinoline-1-oxide-transformed fibroblasts.

The collagen phenotype of a 4-nitroquinoline-1-oxide-transformed line of Syrian hamster embryo fibroblasts, NQT-SHE, was markedly altered from that of normal Syrian hamster embryo cells, which synthesized mainly type I procollagen [pro-alpha 1(I)]2 pro-alpha 2(I). Total collagen synthesis in the transformant was reduced to about 30% of the control level primarily because synthesis of the pro-alpha 1(I) subunit was completely suppressed. The major collagenous products synthesized consisted of two polypeptides, designated as N-33 and N-50, which could be completely separated by precipitation with ammonium sulfate at 33 and 50% saturation, respectively. N-33 migrated similarly to pro-alpha 2(I) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and N-50 migrated slightly more slowly. The collagenous regions of these chains were more sensitive to protease than the analogous region of procollagen I, but alpha-chains could be obtained by digestion for 2 h at 4 degrees C with high ratios of protein:pepsin. Staphylococcus V8 protease and cyanogen bromide peptide maps of N-33 alpha and N-50 alpha chains indicated that the chains were homologous with, but different than, alpha 2(I) chains and that they differed from each other. Considering their similarity to pro-alpha 2(I), it was surprising to find that the N-collagens were secreted to the same extent as was type I procollagen from Syrian hamster embryo cells and that there were no disulfide bonds between N-collagen chains. Intrachain disulfides were present. One possible explanation for the unusual collagen phenotype of NQT-SHE cells is that transformation induced one or more mutations in the pro-alpha 2(I) structural gene while suppression of synthesis of the pro-alpha 1(I) subunit may be due to a mutation in the regulatory region of its gene or in a general regulatory gene.

4-Nitroquinoline-1-oxide↗
PubMed1986

Mechanism for the decreased biosynthesis of cartilage proteoglycan in the scorbutic guinea pig.

Our previous work showed that vitamin C deficiency caused about a 70-80% decrease in the incorporation of [35S]sulfate into proteoglycan of guinea pig costal cartilage, coordinately with a decrease in collagen synthesis (Bird, T. A., Spanheimer, R. G., and Peterkofsky, B. (1986) Arch. Biochem. Biophys. 246, 42-51). We examined the mechanism for decreased proteoglycan synthesis by labeling normal and scorbutic cartilage in vitro with radioactive precursors. Proteoglycan monomers from scorbutic tissue were of a slightly smaller average hydrodynamic size than normal but there was no difference in the size of the glycosaminoglycan chains isolated after papain digestion. The type of glycosaminoglycans synthesized and the degree of sulfation were unaffected as determined by chondroitinase ABC digestion and duel labeling with [35S]sulfate and [3H]glucosamine. Conversion of [3H]glucosamine to [3H]galactosamine also was unimpaired. There was about a 40% decrease in core protein synthesis, measured by [14C]serine incorporation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Nevertheless, decreased incorporation of [35S]sulfate into scorbutic tissue persisted in the presence of p-nitrophenyl-beta-D-xyloside and cycloheximide, which indicated that the site of the scorbutic defect was beyond core protein synthesis and xylosylation. Galactosyltransferase activity in scorbutic cartilage decreased to about one-third the levels in control samples in parallel with the decreases in proteoglycan and collagen synthesis. Our results suggest that the step catalyzed by this enzyme activity, the addition of galactose to xylose prior to chondroitin sulfate chain elongation, is the major site of the scorbutic defect in proteoglycan synthesis. Decreased enzyme activity may be related to increased cortisol levels in scorbutic serum.

Animals↗
PubMed1986

Regulation of collagen synthesis and mRNA levels in articular cartilage of scorbutic guinea pigs.

Previous studies suggested that decreased type I collagen synthesis in calvaria of ascorbate-deficient guinea pigs was correlated with weight loss rather than defective proline hydroxylation. The generality of this correlation was examined in articular cartilage, which synthesizes mainly type II collagen, by measuring collagen synthesis and proline hydroxylation in vitro in tissue from ascorbate-supplemented and scorbutic guinea pigs. Ascorbate concentrations in tissues were almost completely depleted after 1 week of deficiency, but proline hydroxylation remained normal until after approximately 3 weeks, when it had decreased only by 10%. At that point collagen synthesis had decreased to about 50% of the control value. There was little additional effect on proline hydroxylation but collagen synthesis decreased further to 20% of normal. Procollagen mRNA levels in cartilage, as measured by dot-blot hybridization with a type II-specific cDNA probe, were unchanged after 2 weeks of scurvy, which correlated with the lack of effect on collagen synthesis during that period. Thereafter, during the period when collagen synthesis decreased, procollagen mRNA levels decreased to 20% of control values. Refeeding ascorbate to acutely scorbutic animals led to reversal of defective proline hydroxylation within 24 h with a slower increase in collagen synthesis and mRNA levels. Collagen synthesis returned to the normal level after 4 days with no further increase, while mRNA levels continued to increase to 2.7 times the control values after 7 days. Thus the major mechanism for regulation of collagen synthesis in articular cartilage during scurvy and ascorbate repletion occurs independently of the effect on proline hydroxylation and is associated with changes in mRNA levels. The lack of precise coordination between collagen synthesis and mRNA levels during repletion, however, suggests that there may be additional regulation through post-transcriptional mechanisms.

Animals↗
PubMed1986

Coordinate regulation of collagen and proteoglycan synthesis in costal cartilage of scorbutic and acutely fasted, vitamin C-supplemented guinea pigs.

The effects of ascorbic acid deficiency and acute fasting (with ascorbate supplementation) on the synthesis of collagen and proteoglycan in costal cartilages from young guinea pigs was determined by in vitro labeling of these components with radioactive proline and sulfate, respectively. Both parameters were coordinately decreased by the second week on a vitamin C-free diet, with a continued decline to 20-30% of control values by the fourth week. These effects were quite specific, since incorporation of proline into noncollagenous protein was reduced by only 30% after 4 weeks on the deficient diet. The time course of the decrease in collagen and proteoglycan synthesis paralleled the loss of body weight induced by ascorbate deficiency. Hydroxylation of proline in collagen synthesized by scorbutic costal cartilage was reduced to about 60% of normal relatively early, and remained at that level thereafter. Neither collagen nor proteoglycan synthesis was returned to normal by the addition of ascorbate (0.2 mM) to cartilage in vitro. Administration of a single dose of ascorbate to scorbutic guinea pigs increased liver ascorbate and restored proline hydroxylation to normal levels by 24 h, but failed to increase the synthesis of collagen or proteoglycan. Synthesis of both extracellular matrix components was restored to control levels after four daily doses of ascorbate. A 96-h total fast, with ascorbate supplementation, produced rates of weight loss and decreases in the synthesis of these two components similar to those produced by acute scurvy. There was a linear correlation between changes in collagen and proteoglycan synthesis and changes in body weight during acute fasting, scurvy, and its reversal. These results suggest that it is the fasting state induced by ascorbate deficiency, rather than a direct action of the vitamin in either of these two biosynthetic pathways, which is the primary regulatory factor.

Animals↗
PubMed1985

Cysteinyl-cysteine and the microsomal protein from which it is derived act as reducing cofactor for prolyl hydroxylase.

Microsomes from L-929 cells contain a reductant which can replace ascorbate as a cofactor for prolyl hydroxylase. The cofactor was extracted with Triton X-100 and exhibited high and low molecular weight forms on S-300 gel columns. Refiltration or trypsin treatment of high molecular weight cofactor produced additional low molecular weight form. The low molecular weight form was purified by P-2 gel filtration, and Dowex-1 and thin layer chromatography. It is ninhydrin reactive, exhibits reduced and oxidized forms with molecular weights of 240 and 460, respectively, and yielded cystine upon acid hydrolysis. The results suggest that it is a dipeptide, cysteinyl-cysteine, derived from a microsomal protein which is the high molecular weight cofactor.

Animals↗
PubMed1985

A specific decrease in collagen synthesis in acutely fasted, vitamin C-supplemented, guinea pigs.

Weight loss often results from various experimental conditions including scurvy in guinea pigs, where we showed that decreased collagen synthesis was directly related to weight loss, rather than to defective proline hydroxylation (Chojkier, M., Spanheimer, R., and Peterkofsky, B. (1983) J. Clin. Invest. 72, 826-835). In the study described here, this effect was reproduced by acutely fasting normal guinea pigs receiving vitamin C, as determined by measuring collagen and non-collagen protein production after labeling tissues in vitro with [3H]proline. Collagen production (dpm/microgram of DNA) decreased soon after initiating fasting and by 96 h it had reached levels 8-12% of control values. Effects on non-collagen protein were much less severe, so that the percentage of collagen synthesis relative to total protein synthesis was 20-25% of control values after a 96-h fast. These effects were not due to changes in the specific radioactivity of free proline. Refeeding reversed the effects on non-collagen protein production within 24 h, but collagen production did not return to normal until 96 h. The effect of fasting on collagen production was independent of age, sex, ascorbate status, species of animal, and type of connective tissue and also was seen with in vivo labeling. Pulse-chase experiments and analysis of labeled and pre-existing proteins by gel electrophoresis showed no evidence of increased collagen degradation as a result of fasting. Procollagen mRNA was decreased in tissues of fasted animals as determined by cell-free translation and dot-blot hybridization with cDNA probes. In contrast, there was no decrease in translatable mRNAs for non-collagen proteins. These results suggest that loss of nutritional factors other than vitamin C lead to a rapid, specific decrease in collagen synthesis mainly through modulation of mRNA levels.

Age Factors↗
PubMed1985

Cyclic AMP-independent processes mediate Kirsten sarcoma virus-induced changes in collagen production and other properties of cultured cells.

Previous studies suggested that the decreased collagen production observed in Kirsten sarcoma virus (Ki-MSV)-transformed BALB 3T3 cells could be reversed by treating cells with Bt2cAMP. We examined the relationship between intracellular cAMP, collagen production, and other properties in NRK and BALB 3T3 cells transformed by Ki-MSV. Two 3T3 transformants (Ki-3T3-234 and Ki-3T3Cl1) had lower cAMP levels than nontransformed cells. The level in a temperature-sensitive transformant, tsKi-3T3-714, was similar to the level in its parent, 3T3-714, and when it was shifted to a temperature nonpermissive for transformation (40 degrees C), intracellular cAMP did not increase although the growth and morphological properties were normal. The relative rate of collagen production also increased to the normal level. These results indicate that transformation-induced changes were regulated independently of cAMP. Further observations supported this conclusion. Intracellular cAMP in a flat revertant of Ki-3T3Cl1 was lower, rather than higher, than in the transformant, although the relative rate of collagen production was higher. Treatment of Ki-3T3-234 and tsKi-3T3-714 with cholera toxin plus isobutylmethylxanthine increased intracellular cAMP concentrations to 2-20 times the level in untreated cells, levels much higher than in nontransformed 3T3. In spite of this, collagen production was not increased by these agents in tsKi-3T3-714 and it was only partially restored in Ki-3T3-234 relative to the level in the nontransformed cells. In contrast, these agents inhibited growth on a substratum or in soft agar and produced a flattened morphology in both lines. Similarly, collagen production in transformed NRK cells (K-NRK) was only 3% of normal but treatment with Bt2cAMP or cholera toxin plus isobutylmethylxanthine increased production to only one-third the normal level while increasing cAMP to four times the normal level. We conclude that in Ki-MSV-transformed BALB 3T3 cells, changes in cAMP may be secondary effects and not related to maintenance of the transformed phenotype. The high levels of cAMP induced by exogenous agents may act on similar targets as those affected by transformation, but reversal of the transformed phenotype by these agents probably occurs by a different mechanism than that originally used to impose the changes.

1-Methyl-3-isobutylxanthine↗
PubMed1984

Characterization of collagen hydroxylysyl glycosyltransferases as mainly intramembranous microsomal enzymes.

The localization of collagen hydroxylysine galactosyl- and galactosyl-hydroxylysine glucosyltransferases in purified chick embryo bone microsomes was studied by differential solubilization with nonionic detergents. Brij-35 (polyoxyethylene 25-lauryl ether) which selectively releases intracisternal proteins, and Triton X-100, whose specificity varies with its concentration, were used in the presence or absence of high ionic strength NaCl. These methods were used previously to characterize prolyl hydroxylase as intracisternal and lysyl hydroxylase as mainly intramembranous. The distribution of both glycosyltransferases within microsomes was similar to that of lysyl hydroxylase; approximately 70-80% of their activities are intramembranous with the remainder intracisternal. Collagen hydroxylysine glucosyltransferase differed from prolyl and lysyl hydroxylase and the galactosyltransferase in that its activity in vitro was apparently inhibited by membrane vesicles, even in the presence of detergents at concentrations which permeabilize the membrane. Accurate measurement of its activity could be achieved only by its separation from vesicles after detergent treatment. The common location of the major portion of lysyl hydroxylase and the glycosyltransferase activities suggests that they may act as a multienzyme complex to preferentially modify certain lysyl residues in nascent procollagen chains as they traverse the membrane of the endoplasmic reticulum. Since these enzymes do not act on helical collagen, their physical separation from prolyl hydroxylase may ensure that modifications of lysine residues occur prior to formation of hydroxyproline, which stabilizes the helical form.

Animals↗
PubMed1984

Regulation of prolyl hydroxylase activity in L-929 cells by mechanisms unrelated to glycolytic metabolism.

Previous studies showing that prolyl hydroxylase is activated by addition of lactate to logarithmic cultures of L-929 and 3T6 mouse cell lines and that the enzyme normally increases in dense cultures which accumulate lactic acid, have led to the hypothesis that regulation is mediated through glycolytic metabolism. We present evidence that the extent of lactate production plays a minor role in controlling the density-dependent activation of prolyl hydroxylase in L-929 cells. The initial rate of glycolysis in stationary phase cells was lower than that in the logarithmic phase so that intracellular levels of glycolytic intermediates would not be related to activation of the enzyme. Nevertheless, because of increased numbers of cells in stationary phase, higher concentrations of lactate occur in the culture medium. Eliminating accumulation of lactate during the growth of L-929 cells by reducing the normal glucose concentration (5.5 mM) to low-glucose (0.5 mM) did not significantly affect the growth rate and still allowed a 10-fold activation of prolyl hydroxylase when cells reached stationary phase. Growth in normal glucose resulted in only an additional 2-fold increase in specific enzyme activity. These results imply that density-dependent activation of prolyl hydroxylase occurs mainly through a mechanism independent of lactate production. The elevated enzyme activity in dense cells was not decreased when lactate was removed and its production prevented by replacement of high-glucose medium with low-glucose medium for 24 hours. This result differed markedly from the dramatic decrease in enzyme activity observed within several hours after replating stationary phase cells in fresh medium, but at a lower cell density. The conclusion that there are at least two mechanisms regulating prolyl hydroxylase activity in L-929 cells was further supported by the observation that the maximal extent of enzyme activation by ascorbate or lactate in logarithmic phase L-929 cells was much less than the extent of density-dependent activation.

Animals↗
PubMed1983

Specifically decreased collagen biosynthesis in scurvy dissociated from an effect on proline hydroxylation and correlated with body weight loss. In vitro studies in guinea pig calvarial bones.

The question whether ascorbate regulates collagen production solely through its direct role in proline hydroxylation was investigated. Proteins in calvarial bones from control and scorbutic weanling guinea pigs were labeled in short-term cultures with radioactive proline. Proteins were digested with purified bacterial collagenase to distinguish between effects on collagen polypeptide production and hydroxyproline formation. There was a preferential decrease in the absolute rate of collagen biosynthesis beginning after 2 wk of ascorbate deficiency, and this effect was temporally dissociated from decreased proline hydroxylation. There were no significant changes in the absolute rates of collagen degradation or noncollagen protein production. In vitro inhibition of proline hydroxylation in normal bone with alpha, alpha'-dipyridyl did not affect the relative rate of collagen synthesis, further dissociating these functions. Ascorbate added to scorbutic bone cultures reversed defective proline hydroxylation but not defective collagen synthesis, suggesting that the latter was an indirect effect of scurvy. There was a linear correlation between the extent of body weight lost during the 3rd and 4th wk of scurvy and the rate of collagen synthesis in scorbutic bone. This correlation also applied to control animals receiving ascorbate, but with weight loss induced by food restriction. These studies establish for the first time that ascorbate deficiency in guinea pigs leads to a specific decrease in collagen polypeptide synthesis and suggest that this decrease results from the reduced food intake and/or weight-loss characteristic of scurvy.

Animals↗
PubMed1982

Correlation between the rates of aerobic glycolysis and glucose transport, unrelated to neoplastic transformation, in a series of BALB 3T3-derived cell lines.

The relationship between transformation, lactate production, and glucose transport was examined in a series of ten cell lines consisting of subclones of BALB 3T3 A31 cells and viral and chemical transformants of either the subclones or the original A31 line. Comparisons were made over a relatively narrow range of cell densities to minimize changes in the biochemical parameters during growth. A nitroquinoline oxide (NQT-3T3-714) and a temperature-sensitive Kirsten sarcoma virus (tsKi-3T3-714) transformant of subclone 714 exhibited transformed phenotypes with respect to morphology and growth properties, but their rates of lactate production and 2-[3H]deoxy-D-glucose (deoxyglucose) uptake were similar to those of the parent cells. 2- to 5-fold in these transformants, showing that there was no defect in the enzymes of this pathway. At a temperature nonpermissive for transformation of tsKi-3T3-714, lactate production by this line did not decrease relative to the rate of the parent cells. Another transformant, Ki-3T3-234, had a glycolytic rate which was 4 to 5 times greater than that of the low lactate producers while other transformants exhibited intermediate rates, and the rate of a third nontransformed 3T3 A31 subclone, K-1-1, was comparable to the rate of Ki-3T3-234. The rates of [3H]deoxyglucose uptake by this series of cells were closely proportional to their glycolytic rates rather than to their state of transformation. Increasing glycolysis by oligomycin or dinitrophenol treatment, however, did not cause a concomitant increase in sugar uptake. Neither glycolysis nor deoxyglucose uptake in the high-lactate producer (Ki-3T3-234) was inhibited by ouabain, suggesting that Na+-K+-adenosinetriphosphatase is not a regulatory of these functions in 3T3 cells. In 3T3-derived cells, it appears that the rates of glycolysis and glucose uptake may be regulated in tandem under some conditions and that neither process is an obligatory consequence of neoplastic transformation

4-Nitroquinoline-1-oxide↗
PubMed1981

Mechanisms of Kirsten murine sarcoma virus transformation-induced changes in the collagen phenotype and synthetic rate of BALB 3T3 cells.

Specific viral transformation rather than cell selection can explain the previously observed increase in the proportion of type III procollagen compared to type I procollagen in BALB 3T3 cells transformed by Kirsten murine sarcoma virus (Ki-MSV). Two subclones of BALB 3T3 A31 were productively infected with with a temperature-sensitive Ki-MSV in the presence of helper murine leukemia virus (MLV), resulting in virtually complete transformation of cultures and eliminating selection of transformed foci. Analysis of radioactive collagen, derived from procollagen by pepsin treatment, showed that both of the tsKi-MSV/MLV-transformed subclones contained a 4-fold greater proportion of type III procollagen than did control MLV-infected cultures. A nonproducer derivative exhibited an even greater change (10-fold), indicating that viral replication was irrelevant. After 48 hr at a nonpermissive temperature, tsKi-MSV-transformed cells retained a high proportion of type III procollagen, suggesting that either this change is not induced by src protein or else there is a slowly reversible or irreversible step involved. Alternatively, type III procollagen mRNA may be long lived. In contrast, the relative rate of procollagen synthesis in transformed cells was clearly regulated by src protein. Translation of mRNA from cells preincubated at permissive or nonpermissive temperatures revealed that the decreased relative rate can be explained by a simultaneous small decrease in the level of procollagen mRNA and a large increase in mRNA for noncollagen proteins.

Animals↗