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Biomedical subjects

S Kit

Publications and source records attributed to S Kit.

At least 91 records · Page 5Linked to original sources
PubMed1976

Thymidine kinase, DNA synthesis and cancer.

A resume has been presented of some recent investigations which show that DNA synthesis can be initiated in many types of quiescent animal cells by external stimuli, by introducing a quiescent nucleus into the cytoplasm of a proliferating cell, or by a virus infection. The components of the DNA replication apparatus are described. It is shown that deoxyribonucleoside triphosphate pools increase substantially in animal cells at the time DNA synthesis is initiated due to the enhanced activities of enzymes functioning in nucleotide synthesis. Especially striking is the increase of thymidine kinase activity, indicating that this enzyme may be a useful marker of the shift from the quiescent to the replicative state. The thymidine kinase isozymes of vertebrate cells have been characterized. Thymidine kinase F, which is found principally in the cytosol, is the isozyme that increases when G1 (Go) phase cells are stimulated or infected with oncogenic viruses. Chick cytosol thymidine kinase F can also be reactivated by introducing differentiated chick erythrocyte nuclei into the cytoplasm of enzyme-deficient LM (TK-) mouse cells. Furthermore, herpesviruses code for distinctive, virus-specific thymidine kinase isozymes, so that another way to transform thymidine kinase-deficient LM TK-) cells to kinase-positive cells is by infecting them with UV-irradiated herpes simplex viruses. The experiments on the activation of DNA synthesis and thymidine kinase F activity have been discussed in the context of the proliferative activity in vivo and the immortalization in culture of neoplastic cells. These experiments suggest that genes determining cell cycle proteins are readily accessible to transcription and translation in essentially all nucleated cells. The tendency of transformed cells to become multinucleated after cytochaliasin B treatment also suggests that one important difference between malignant cells and most normal cells may be the ability of malignant cells to 'stockpile' the proteins (and/or their messenger RNAs) of the DNA replicative apparatus and to maintain the 'stockpiles' in progeny cells.

Animals↗
PubMed1976

Formation of thymidine kinase and deoxycytidylate deaminase in synchronized cultures of chinese hamster cells temperature-sensitive for DNA synthesis.

Cytosol thymidine kinase (TK) and deoxycytidylate (dCMP) deaminase formation was investigated in synchronized cultures of K12 Chinese hamster cells which have a temperature-sensitive lesion affecting the initiation of DNA synthesis. Enzyme formation was found to be cycloheximide-sensitive and also temperature-dependent. Beginning at about six hours after addition of medium with 10% calf serum to serum-depleted K12 cultures, cytosol TK and dCMP deaminase activities increased when the cultures were incubated at 36.5 degrees but not at 40.5 degrees. When cultures were shifted from 36.5 degrees to 40.5 degrees at 4,6, or 8 hours after serum addition, TK activity continued to increase, though not to the level observed at ten hours in cultures maintained at 36.5 degrees. Actinomycin D addition at the time of serum reversal or four hours later blocked the TK increase normally observed at the permissive temperature at ten hours. However, when actinomycin D addition was delayed for six or eight hours after serum addition, the increase in TK measured at ten hours resembled that observed in the temperature shift-up experiments. The results provide evidence that the mutation in K12 Chinese hamster cells most likely blocks the progression through G1 into S and suggest that transcription or post-transcriptional processing required for TK formation is affected.

Blood↗
PubMed1976

Reactivation of chick erythrocyte nuclei in heterokaryons with temperature-sensitive Chinese hamster cells.

Chinese hamster cell line K12 is temperature-sensitive for the initiation of DNA synthesis. K12 cells synchronized by serum deprivation were collected in early G1(G0). Heterokaryons were formed by fusing chick erythrocytes with serum-starved K12 cells through the use of UV-irradiated Sendai virus. At the permissive temperature (36.5 degrees C), erythrocyte nuclei in heterokaryons enlarged, the chromatin dispersed, and erythrocyte nuclei synthesized DNA at about the same time as the K12 nuclei. At the restrictive temperature (41 degrees C), erythrocyte nuclei enlarged, but neither erythrocyte nor K12 nuclei initiated DNA synthesis. When erythrocyte nuclei were fused with Wg-1A cells, the wild-type parent for ts K12 cells, both kinds of nuclei synthesized DNA at 36.5 degrees C and 41 degrees C. Activation of erythrocyte nuclei was inefficient in heterokaryons incubated in low-serum medium. The results indicate that serum factors and a cellular function defined by the K12 mutation are required for activation of chick erythrocyte nuclear DNA synthesis.

Blood↗
PubMed1975

Analyses of deoxycytidylate deaminase molecular forms in human-mouse and monkey-mouse somatic cell hybrids.

Disc polyacrylamide gel electrophoresis (disc PAGE) analyses have revealed that mouse, human, and monkey cytosal deoxycytidylate (dCMP) deaminases differ in electrophoretic mobility, so that mixtures of mouse and human, mouse and monkey and human and monkey enzymes can be separated. To learn whether the genes for dCMP deaminase and thymidine (dT) kinase are genetically linked, disc PAGE analyses of cytosol fractions from human-mouse and monkey-mouse somatic cell hybrids were carried out. The interspecific somatic cell hybrids were derived from the fusion of cytosol dT kinase deficient mouse cells with cytosol dT kinase-positive human and monkey cells: they contained mostly mouse chromosomes and a few primate chromosomes, including the determinant for primate cytosol dT kinase. The disc PAGE analyses demonstrated that the human-mouse and monkey-mouse somatic cell hybrids contained a dCMP deaminase activity with an electrophoretic mobility characteristic of mouse dCMP deaminase. Enzymes with electrophoretic mobilities characteristic of human and monkey dCMP deaminases were not demonstrable. these findings suggest that primate cytosol dT kinase and dCMP deaminase are coded on different chromosomes, or that the formation in hybird cells of an active primate dCMP deaminase is suppressed. Chick-mouse somatic cell hybrids containing chick but not mouse cytosol dT kinase were also analyzed, but it was not possible to establish whether the enzyme was of murine or avian origin because of the similarity in electrophoretic mobility between the chick and mouse enzymes. Human and mouse cells contained low levels of mitochondrial dCMP deaminase activity. In contrast to dT kinase isozymes, however, mitochondrial and cytosol dCMP deaminases were electrophoretically indistinguishable.

Aminohydrolases↗
PubMed1975

Mitochondrial and herpesvirus-specific deoxypyrimidine kinases.

To characterize and compare the thymidine (TdR) and deoxycytidine (CdR) kinase isozymes of uninfected and herpesvirus-infected cells: (i) the subcellular distribution of the isozymes has been studied; (ii) a specific assay for CdR kinase has been devised; (iii) the TdR kinase isozymes have been partially purified; and (iv) the purified enzymes have been analyzed by disc polyacrylamide gel electrophoresis, isoelectric focusing, and glycerol gradient centrifugation and by substrate competition and dCTP inhibition studies. The results indicate that there are interesting individual differences with respect to nucleoside acceptor specificity between the cytosol and mitochondrial pyrimidine deoxyribonucleoside kinases of uninfected cells and between the enzymes induced by different herpesviruses. In the cytosol of uninfected mouse, chicken, and owl monkey kidney cells, two different proteins, TdR kinase F and CdR kinase 2, catalyze the phosphorylations of TdR and CdR, respectively. TdR kinase F does not phosphorylate CdR, nor does CdR kinase 2 phosphorylate TdR. A second TdR kinase isozyme present in HeLa(BU25) mitochondria (TdR kinase B) also lacks CdR phosphorylating activity. In contrast, a genetically distinctive deoxypyrimidine kinase (TdR kinase A) of mouse, human, and chick mitochondria catalyzes the phosphorylation of both TdR and CdT. Three herpesviruses, marmoset herpesvirus and herpes simplex virus types 1 and 2, induce in the cytosol fraction of LM(TK-) mouse cells isozymes which share common properties with mitochondrial TdR kinase A, including the ability to catalyze the phosphorylation of both TdR and CdR. However, the herpesvirus-induced deoxypyrimidine kinases differ from mitochondrial TdR kinase A with respect to sedimentation coefficient, sensitivity to dCTP inhibition, and antigenic determinants. The herpesvirus-specific and the mitochondrial deoxypyrimidine kinases exhibit a preference for TdR over CdR as nucleoside acceptor. Pseudorabies virus and herpesvirus of turkeys induce cytosol TdR kinases resembling the other herpesvirus-induced TdR kinases in several properties, but like cellular TdR kinase F, the pseudorabies virus and herpesvirus of turkeys TdR kinases lack detectable CdR phosphorylating activities. Finally, a marmoset herpesvirus nutant resistant to bromodeoxyuridine, equine herpesvirus type 1, and Herpesvirus aotus induces neither TdR nor CdR phosphorylating enzymes during productive infections.

Cell Line↗
PubMed1974

Genetic control of mitochondrial thymidine kinase in human-mouse and monkey-mouse somatic cell hybrids.

Distinctive thymidine (dT) kinase molecular forms are present in mouse, human, and monkey mitochondria. Disk polyacrylamide gel electrophoresis (disk PAGE) analyses have shown that the mitochondrial-specific dT kinases differ from cytosol dT kinases in relative electrophoretic mobilities (Rm). Furthermore, the mouse mitochondrial dT kinase differs in Rm value from primate mitochondrial dT kinases. The mouse and primate cytosol dT kinases can also be distinguished. Disk PAGE analyses have been carried out on the cytosol and mitochondrial dT kinases of human-mouse (WIL-8) and monkey-mouse (mK.CV(III)) somatic cell hybrids in order to learn whether the mitochondria of the hybrid cells contained murine mitochondrial-specific, primate mitochondrial-specific, or both dT kinases. WIL-8 cells were derived from cytosol dT kinase-negative, mitochondrial dT kinase-positive mouse fibro blasts and from cytosol dT kinase-positive, mitochondrial dT kinase-positive human embryonic lung cells; they contained mostly mouse chromosomes and a few human chromosomes, including the determinant for human cytosol dT kinase. The mK.CV(III) cells were derived from cytosol dT kinase-negative, mitochondrial dT kinase-positive mouse kidney cells and from cytosol dT kinase-positive, mitochondrial dT kinase-positive monkey kidney cells; they contained mostly mouse chromosomes and a few monkey chromosomes, including the determinant for monkey cytosol dT kinase. Disk PAGE analyses demonstrated that the mitochondria of human-mouse and monkey-mouse somatic cell hybrids contained the mouse-specific mitochondrial dT kinase but not the human- or monkey-specific mitochondrial dT kinase. These findings suggest that primate cytosol and mitochondrial thymidine kinase genes are coded on different chromosomes.

Animals↗