摘要:

AbstractA temperature‐sensitive mutant of Chinese hamster cells is described which has two interesting properties: (1) it is a cell cycle mutant and (2) glycoprotein synthesis appears to be affected at the non‐permissive temperature (40°C). Synchronized cells shifted to 40°C in the beginning of their G1phase do not incorporate [3H]‐thymidine into DNA during the expected S‐phase, but once DNA synthesis has been initiated (∼ 10 hours after termination of serum starvation) a shift to 40°C no longer leads to an arrest of DNA synthesis. Flow microfluorimetric analysis of DNA content/cell supports this conclusion and indicates that a majority of cells become arrested in the G1phase of the cell cycle when a non‐synchronized population of cells is transferred to 40°C. Apparently at all times in the cell cycle there is a drastic reduction of incorporation of labeled sugars (particularly fucose) into glycoproteins. The uptake of fucose and its conversion to GDP‐fucose appears to be normal at 40°C. Chromatographic analysis indicates that all classes of glycoproteins are affected, and we do not find any evidence for partially completed oligosaccharides at 40°C. Overall protein synthesis is not reduced at the nonpermissive temperature during the time interval under consideration and the number of polysomes attached to membranes (RER) is also normal at 40°C. This suggests that the defect is at an early step in the synthesis or regulation of synthesis of glycoproteins. The mutation is a recessive mutation in hybrid cells and mutagen induced revertants can be obtained which grow normally at 40°C and in which glycoprotein synthesis at 40°C is restored to no

ISSN:0021-9541    

DOI:10.1002/jcp.1040900202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractThe thermodynamic parameters ΔH°, ΔF°, ΔS° and the pH dependence of the interaction between steroids and the glucose carrier in human erythrocytes have been studied. The results indicate that, according to the structure of the steroids, hydrophilic as well as hydrophobic bonds can be involved in the association. For the binding of the C‐21‐steroids to the carrier the polarity rule has been observed to be valid. On the basis of the thermodynamic parameters it has been found that in the association process of the steroids with the carrier, simultaneous randomization of ordered water molecules is

ISSN:0021-9541    

DOI:10.1002/jcp.1040900203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractWhenParamecium tetraureliain log phase growth is treated with 4% dimethyl sulphoxide (DMSO) for five minutes the amount of polyribosomes is reduced 3‐ to 4‐fold while there is a corresponding increase in 80s ribosomal material. Reducing the concentration of DMSO to 1% allows immediate reversal of the condition.Parameciumpolyribosomes subjected to 4% DMSO either in whole cell homogenates or during purification through sucrose density gradients appear unaffected while cycloheximide at concentrations up to 100 μg/ml did not prevent DMSO from exerting its effect in vivo.Analyses of14C amino acid incorporation experiments indicated a strict correspondence between the effect of DMSO on polyribosomes and overall protein synthesis. The reduction of acid precipitable radioactivity in the polyribosomal region after DMSO treatment was associated with a corresponding increase in radioactivity in the 80s region. There was no comparable increase in the acid precipitable radioactivity in the soluble fraction.The overall results of the study suggest that DMSO acts on polyribosomes indirectly through some unknown primary reaction with cell constituents, and that the mode of action is such as to cause the release of ribosomes from messenger RNA (mRNA) rather than to prevent initiation of the ribosome‐mRNA complex. Our data suggest that the effect may be selective.Finally, it is of interest that high concentrations of DMSO (above 8%) appear to have the opposite effect of lower concentrations of DMSO, i.e., they appear to “freeze” the ribosom

ISSN:0021-9541    

DOI:10.1002/jcp.1040900204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractSomatic cell hybrids of mouse L‐cells with rat HTC cells were studied for their growth properties in vitro and tumorigenicity in vivo. Three hybrid clones were selected for detailed study. All were delayed in tumor formation innudemice compared with the parent lines despite their varied growth properties in vitro.One clone was sensitive to density dependent inhibition of growth (DDIG) and had a relatively low saturation density. A second clone was not sensitive to DDIG and had a higher saturation density. The third clone had atypical morphology, was insensitive to DDIG, and had a relatively high saturation density. All of the clones studied produced colonies suspended in agarose gel which were much smaller than those of the parents incubated for the same period of time. Only the pattern of growth in agarose gel corresponded to the delayed tumor formation in vivo of the hybrids. Sensitivity to DDIG and saturation density were not consistent with tumor growth.The hybrid clone that was sensitive to DDIG was the only one of the three that had a nearly complete set of chromosomes derived from each of the parents. The chromosome numbers of all three clones were unchanged after growth in agarose or as tumors in thenudemic

ISSN:0021-9541    

DOI:10.1002/jcp.1040900205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractPrevious studies have shown that initiation of proliferation of density‐inhibited fibroblasts by fresh serum is accompanied by a rapid increase in phosphate uptake. This increase might be a key event in the initiation of DNA synthesis. The present studies examined this possibility.Mouse 3T3, secondary chick embryo, or human diploid foreskin cultures were grown to quiescence in medium containing varying levels of serum. When proliferation of the cultures was initiated by addition of fresh serum, the changes in phosphate uptake were inversely related to the final increases in cell number. Additional experiments showed that the change in phosphate uptake following serum addition was determined by the level of phosphate uptake prior to serum addition.Addition of dexamethasone to quiescent 3T3 cultures caused them to proliferate but did not increase phosphate uptake. Similarly, trypsin or insulin stimulated proliferation of quiescent secondary chick embryo cultures, but caused little or no change in phosphate uptake.Quiescent 3T3 cultures switched to medium containing fresh serum and reduced levels of phosphate showed a decrease in both phosphate uptake and intracellular phosphate pool size. Cell proliferation in these cultures, however, was stimulated to the same degree as cultures switched to medium containing fresh serum and the normal amount of phosphate. In addition, quiescent secondary chick embryo cultures switched to medium containing fresh serum and no phosphate showed a decrease in the intracellular phosphate pool size. Thymidine incorporation and final cell number in these cultures, however, was stimulated to the same or higher degree than in cultures switched to medium containing fresh serum and the normal amount of phosphate. These results demonstrate that the rapid increase in phosphate uptake following addition of fresh serum to quiescent fibroblasts is not a necessary event for the initiation of proliferatio

ISSN:0021-9541    

DOI:10.1002/jcp.1040900206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractIn resting, non‐growing human diploid fibroblasts the amount of rRNA is reduced 1.8‐fold, cytoplasmic polysomes are disaggregated, and the level of poly‐A RNA (mRNA) is reduced 1.8‐fold in relation to growing cells. The distribution of poly‐A RNA is altered in resting, non‐growing cells so that an average of 64% of the total cytoplasmic poly‐A RNA sediments along with particles lighter than 80S (prepolysomal) in sucrose density gradients. By comparison, in growing cells only 30% of the cytoplasmic poly‐A RNA sediments in the prepolysomal region. In SDS sucrose gradients, the sedimentation profile of the prepolysomal poly‐A RNA from resting cells resembles that of polysomal poly‐A RNA from those cells. In contrast, the average size of prepolysomal poly‐A RNA from growing cells is much smaller than that of the polysomal poly‐A RNA from those cells. These data are compatible with the possibility that resting cell prepolysomal poly‐A is untranslated mRNA. Also consistent with this interpretation are experiments which demonstrate that one‐quarter to one‐third of the prepolysomal poly‐A RNA of resting cells is recruited into polysomes i

ISSN:0021-9541    

DOI:10.1002/jcp.1040900207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractEndothelial cells from bovine aorta and human umbilical vein and fibroblasts from human foreskin were cultured and subsequently evaluated for ability to metabolize serotonin (5‐HT) to 5‐hydroxyindoleacetic acid (5‐HIAA). Cells were incubated for three hours with 4 × 10−6M [14C] 5‐HT creatinine sulfate. [14C] 5‐HIAA was separated from labeled 5‐HT by column chromatography and measured by scintillation counting. Production of 5‐HIAA by bovine aorta cells was 39.0 ± 7.5 (S.E.M., n = 6) nmoles per 109cells per hour. Production of 5‐HIAA was markedly inhibited by the presence of 10−4M iproniazid (an inhibitor of monoamine oxidase) or 10−4M imipramine (an inhibitor of amine transport). 5‐HIAA was the only product of 5‐HT metabolism detected by thin layer chromatography. Production of 5‐HIAA by human umbilical vein endothelial cells was 5.4 ± 2.0 nmoles per 109cells per hour (n = 5) and by human foreskin fibroblasts was 3.9 ± 1.4 nmoles per 109cells per hour (n = 5). The results obtained during incubation in the presence and absence of inhibitors indicate that bovine aorta endothelial cells maintained in tissue culture are able to transport serotonin with subsequent production of 5‐HIAA. By contrast, human umbilical vein endothelial cells and fibroblasts exhibited relatively low ra

ISSN:0021-9541    

DOI:10.1002/jcp.1040900208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractCultured mouse and human cells were arrested in their growth by artificially depriving them of phosphate. The quiescent cells could be stimulated to synthesize DNA and to divide by addition to the growth medium of insulin, dialyzed serum and/or the full concentration of phosphate. In order to gain insight into mechanisms by which insulin and serum stimulate growth, the inhibitory effects of antimitotic agents were examined. Of the inhibitors tested, vinblastine and cytocalasin B abolished the growth promoting activity of insulin, while colchicine inhibited the activity of both serum and insulin. The present results suggest that insulin‐stimulated growth is mediated by a different pathway than serum‐stimulated growth and is sensitive to mechanisms that occur at various times prior to insulin addit

ISSN:0021-9541    

DOI:10.1002/jcp.1040900209

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractErythropoietin (epo) added to liquid cultures of mouse bone marrow cells affected both the numbers of spleen colony‐forming cells (CFU) in the cultures and the types of spleen colonies formed from these cells in irradiated hosts. Epo caused an increase in the numbers of CFU detected in cultures on the second day; this effect persisted through day 10, with the maximal increase occurring on the seventh day. The magnitude of the rise on day 7 was proportional to the amount of epo added. The increase in spleen colonies found with cells cultured in the presence of epo was due solely to erythroid colonies. After seven days in culture without epo, there was a peak of cells that formed non‐erythroid colonies. This peak did not appear when the cells were cultured in the presence of

ISSN:0021-9541    

DOI:10.1002/jcp.1040900210

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY

摘要:

AbstractA protease fromTetrahymena pyriformisinactivated eight of nine commercially available enzymes tested, including lactate dehydrogenase, isocitrate dehydrogenase (TPN‐specific), glucose‐6 phosphate dehydrogenase, D‐amino acid oxidase, fumarase, pyruvate kinase, hexokinase, and citrate synthase. Urate oxidase was not inactivated. Inactivation occurred at neutral pH, was prevented by inhibitors of the protease, and followed first order kinetics. In those cases tested, inactivation was enhanced by mercaptoethanol. Most of the enzyme‐inactivating activity was due to a protease of molecular weight 25,000 that eluted from DEAE‐Sephadex at 0.3MKCl. A second protease of this molecular weight, which was not retained by the gel, inactivated only isocitrate dehydrogenase and D‐amino acid oxidase. These two proteases could also be distinguished by temperature and inhibitor sensitivity. Two other protease peaks obtained by DEAE‐Sephadex chromatography had little or no enzyme inactivating activity, while another attacked only D‐amino acid oxidase.At least six of the enzymes could be protected from proteolytic inactivation by various ligands. Isocitrate dehydrogenase was protected by isocitrate, TPN, or TPNH, glucose‐6‐dehydrogenase by glucose‐6‐P or TPN, pyruvate kinase by phosphoenolypyruvate or ADP, hexokinase by glucose, and fumarase by a mixture of fumarate and malate. Lactate dehydrogenase was not protected by either of its substrates or coenzymes. Citrate synthase was probably protected by oxalacetate.Our data suggest that the protease or proteases discussed here may participate in the inactivation or degradation of at least some enzymes inTetrahymena. Since the inactivation occurs at neutral pH, this process could be regulated by variations in the cellular levels of substrates, coenzymes, or allosteric regulators resulting from changes in growth conditions or growth state. Such a mechanism would permit the selective retention of enzymes of metabo

ISSN:0021-9541    

DOI:10.1002/jcp.1040900211

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1977

数据来源: WILEY