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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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.
Mol Cell Biochem 1976 Jun 15
PMID:Thymidine kinase, DNA synthesis and cancer. 94 May 49

Four ubiquitin mRNA size classes were found to be differentially regulated in mesophyll protoplast-derived cultures of Nicotiana sylvestris. Three mRNA families of 1.9, 1.6 and 1.35 kb were expressed as soon as protoplasts were isolated. The 1.9 and 1.6 kb size classes were transiently expressed during the first hours of culture, whereas the level of expression of the 1.35 kb size class was maintained as long as cells kept dividing. A 0.7 kb mRNA size class started to be expressed just before the first divisions were observed. cDNAs corresponding to each of these families were isolated from a 6-h-old protoplast cDNA library and characterized. The 1.9, 1.6 and 1.35 kb mRNAs thus encode 7- or more, 6- and 5-mers, respectively, of ubiquitin whereas the 0.7 kb mRNAs encode a monomer of ubiquitin fused to a carboxyl extension protein of 52 amino acids. The expression of ubiquitin genes was studied, using probes specific for each of these transcript families, during protoplast culture and, for comparison, after various stresses including heat shock, HgCl2 treatment, a viral infection giving rise to a hypersensitive reaction, and an Agrobacterium tumefaciens infection which resulted in tumour formation. The 1.9 and 1.6 kb mRNA size classes were found to be stress-regulated, the 0.7 kb mRNA size class developmentally regulated and the 1.35 kb size class both stress- and developmentally regulated.
Plant Mol Biol 1992 Dec
PMID:Ubiquitin genes are differentially regulated in protoplast-derived cultures of Nicotiana sylvestris and in response to various stresses. 128 39

1. We have demonstrated previously (Harrowe et al., 1990), using a lymphoblastoid cell line that constitutively expresses the substance P receptor (SPR) (Payan et al., 1984, 1986), that this receptor may facilitate measles virus (MV) fusion with these cells. In order to test this hypothesis further, a stable cell line transfected with SPR cDNA has been established, and various stages of MV infection in SPR positive and negative cells compared. 2. Jurkat cells, a human T-lymphoblastoid cell line, were transfected with a cDNA clone encoding the SPR. Cells transfected with only the plasmid were used as controls. Jurkat cells and Jurkat vector control cells (J-vo) failed to demonstrate any detectable 125I-SP binding, whereas a clonally selected population of cells transfected with SPR cDNA (J-SPR) expressed about 50,000 receptors/cell (Sudduth-Klinger et al., 1992). 3. Using the J-vo- and J-SPR-transfected cell lines, the following experiments were conducted to investigate the effect of SPR expression on MV infection. To determine if MV would preferentially attach to J-SPR as compared to J-vo, we absorbed virus to cells at 37 degrees C for various times and measured bound MV using a fluorescence activated cell sorter (FACS). Using this approach, we found that MV bound to a greater degree to J-SPR compared with J-vo. In addition to equilibrium being reached faster for J-SPR, the total amount of bound MV was higher on J-SPR. The effect was greater at lower MOIs, suggesting that there existed multiple binding sites for MV on these cells and that the affinity is higher for those cells expressing the SPR. 4. Since binding does not necessitate a successful viral infection, we needed to know if this difference in binding reflected a difference in infection. This was demonstrated by showing an approximate twofold increase in infected cells after a 2-hr binding period with J-SPR as compared to J-vo at an MOI of 1 in an infectious cell-center assay. Moreover, when both cells types were subjected to continuous infection in culture, J-SPR-infected cells produced a seven- to ninefold increase in measles viral titer in 24 hr as compared with J-vo. The observed increase in viral titer may have resulted in more of the J-SPR cells binding virus, as indicated by our binding and infectious cell-center data, or alternatively, the virus might have entered the J-SPR cells faster and begun replication before the J-vo-infected cells.(ABSTRACT TRUNCATED AT 400 WORDS)
Cell Mol Neurobiol 1992 Oct
PMID:Measles virus-substance P receptor interaction: Jurkat lymphocytes transfected with substance P receptor cDNA enhance measles virus fusion and replication. 128 55

Tobacco plants transformed with the sequence encoding the 54-kDa putative replicase protein of tobacco mosaic virus were resistant to systemic virus disease (D. B. Golemboski, G. P. Lomonossoff, and M. Zaitlin, Proc. Natl. Acad. Sci. USA 87:6311-6315, 1990). Resistance was due to a marked suppression of virus replication at the site of inoculation (J. P. Carr and M. Zaitlin, Mol. Plant-Microbe Interact. 4:579-585, 1991). Although RNA transcripts encoding the 54-kDa protein were present in resistant plants, the 54-kDa protein itself was not observed in vivo. We wished to assess the relative importance of the 54-kDa protein versus its RNA in mediating resistance. Further attempts to detect the 54-kDa protein in plant tissues were unsuccessful; therefore, an indirect approach was taken using a protoplast-based transient gene expression system. Electroporation of protoplasts with plasmids capable of expressing the wild-type 54-kDa protein gene sequence or a mutant lacking the first AUG initiation codon of the 54-kDa open reading frame and encoding a slightly truncated protein reduced virus replication in protoplasts. In contrast, a frameshift mutant that was capable of directing synthesis of a protein only 20% the size of the 54-kDa protein, did not produce resistance in protoplasts. These results show that expression of the 54-kDa protein gene sequence at the RNA level alone is insufficient for resistance, and they implicate the 54-kDa protein itself in mediating this resistance phenomenon.
Mol Plant Microbe Interact
PMID:Resistance to tobacco mosaic virus induced by the 54-kDa gene sequence requires expression of the 54-kDa protein. 128 44

Various polyoxometalates proved inhibitory to the replication of a number of enveloped DNA and RNA viruses, i.e., herpesviruses (herpes simplex and cytomegalo), togaviruses (Sindbis), paramyxoviruses (respiratory syncytial), rhabdoviruses (vesicular stomatitis), arenaviruses (Junin and Tacaribe), and retroviruses [human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2), simian immunodeficiency virus, and murine sarcoma virus]. The most potent compounds, i.e., JM1590 [K13[Ce(SiW11O39)2]. 26H2O] and JM2766 [K6[BGa(H2O)W11O39]. 15H2O], inhibited HIV-1 and simian immunodeficiency virus at concentrations as low as 0.008-0.8 microM. The polyoxometalates also inhibited giant cell formation in co-cultures of HIV-infected HUT-78 cells and uninfected MOLT-4 cells. Studies designed to unravel the mechanism of action of these compounds revealed that they inhibit the reverse transcriptase activity associated with HIV. The polyoxometalates also proved inhibitory to the binding of HIV-1 virions to the cells. From "time of addition" experiments, whereby the polyoxometalates were added at different times after virus infection, their mechanism of anti-HIV action could be attributed to inhibition of virus-cell binding. There was a good correlation (r = 0.84) between the inhibitory effects of the compounds on HIV-1-induced cytopathicity and their inhibitory effects on syncytium formation and a close correlation (r = 0.902) between their inhibitory effects on syncytium formation and their interaction with gp120, whereas there was no correlation between their anti-HIV-1 activity and their inhibitory effects on HIV-1 reverse transcriptase. In flow cytometric studies, the compounds did not interfere with the binding of OKT4A/Leu-3a monoclonal antibody to the CD4 receptor of uninfected cells, but they inhibited binding of anti-gp120 monoclonal antibody to HIV-1-infected cells. Thus, the binding of the polyoxometalates to the viral envelope glycoprotein gp120 is responsible for their anti-HIV activity.
Mol Pharmacol 1992 Dec
PMID:Mechanism of anti-human immunodeficiency virus action of polyoxometalates, a class of broad-spectrum antiviral agents. 128 64

(S)-1-[3-Hydroxy-2-(phosphonylmethoxy)propyl]cytosine (HPMPC) is an antiviral phosphonate nucleotide analogue that displays activity against a range of herpesviruses. Anion exchange high performance liquid chromatography analysis of the 60% methanol extract from [14C]HPMPC-treated cells reveals the formation of three major metabolites. Two of these were identified as phosphorylated forms of HPMPC, HPMPC phosphate, and HPMPC diphosphate, by liberation of HPMPC upon acid digestion and coelution with synthetic standards on high performance liquid chromatography. The third metabolite, which is resistant to alkaline phosphatase cleavage but sensitive to phosphodiesterase, is proposed to be an HPMPC phosphate adduct. In herpes simplex virus-1-infected cells the same three metabolites are detected, at concentrations comparable to those in uninfected cells. When HPMPC is removed from the medium, the concentrations of the metabolites in cells decrease slowly, with half-lives of approximately 6, 17, and 48 hr for HPMPC phosphate, HPMPC diphosphate, and the HPMPC phosphate adduct, respectively. HPMPC diphosphate inhibits herpes simplex virus-1 and -2 DNA polymerases with a lower Ki than that for DNA polymerase alpha, and enzyme inhibition is competitive in each case. The formation and the persistence of HPMPC phosphates in cells and the selective inhibition of viral DNA polymerases by HPMPC diphosphate can explain why cells pretreated with HPMPC remain refractory to viral infection even long after HPMPC is removed from the medium.
Mol Pharmacol 1992 Jan
PMID:Intracellular metabolism of the antiherpes agent (S)-1-[3-hydroxy-2-(phosphonylmethoxy)propyl]cytosine. 131 Jan 43

A series of genetic deletions based partly on two RNA secondary structure models (M. A. Skinner, V. R. Racaniello, G. Dunn, J. Cooper, P. D. Minor, and J. W. Almond, J. Mol. Biol. 207:379-392, 1989; E. V. Pilipenko, V. M. Blinov, L. I. Romanova, A. N. Sinyakov, S. V. Maslova, and V. I. Agol, Virology 168:201-209, 1989) was made in the cDNA encoding the 5' noncoding region (5' NCR) of the poliovirus genome in order to study the sequences that direct the internal entry of ribosomes. The modified cDNAs were placed between two open reading frames in a single transcriptional unit and used to transfect cells in culture. Internal entry of ribosomes was detected by measuring translation from the second open reading frame in the bicistronic mRNA. When assayed alone, a large proportion of the poliovirus 5' NCR superstructure including several well-defined stem-loops was required for ribosome entry and efficient translation. However, in cells cotransfected with a complete infectious poliovirus cDNA, the requirement for the stem-loops in this large superstructure was reduced. The results suggest that virus infection modifies the cellular translational machinery, so that shortened forms of the 5' NCR are sufficient for cap-independent translation, and that the internal entry of ribosomes occurs by two distinct modes during the virus replication cycle.
 ...
PMID:Intracellular modifications induced by poliovirus reduce the requirement for structural motifs in the 5' noncoding region of the genome involved in internal initiation of protein synthesis. 131 Jul 72

We have studied the initiation of transcription in vitro by RNA polymerase II on simian virus 40 (SV40) minichromosomal templates isolated from infected cells. The efficiency and pattern of transcription from the chromatin templates were compared with those from viral DNA templates by using two in vitro transcription systems, either HeLa whole-cell extract or basal transcription factors, RNA polymerase II, and one of two SV40 promoter-binding transcription factors, LSF and Sp1. Dramatic increases in numbers of transcripts upon addition of transcription extract and different patterns of usage of the multiple SV40 initiation sites upon addition of Sp1 versus LSF strongly suggested that transcripts were being initiated from the minichromosomal templates in vitro. That the majority of transcripts from the minichromosomes were due to initiation de novo was demonstrated by the efficient transcription observed in the presence of alpha-amanitin, which inhibited minichromosome-associated RNA polymerase II, and an alpha-amanitin-resistant RNA polymerase II, which initiated transcription in vitro. The pattern of transcription from the SV40 late and early promoters on the minichromosomal templates was similar to the in vivo pattern of transcription during the late stages of viral infection and was distinct from the pattern of transcription generated from viral DNA in vitro. In particular, the late promoter of the minichromosomal templates was transcribed with high efficiency, similar to viral DNA templates, while the early-early promoter of the minichromosomal templates was inhibited 10- to 15-fold. Finally, the number of minichromosomes competent to initiate transcription in vitro exceeded the amount actively being transcribed in vivo.
Mol Cell Biol 1992 Apr
PMID:In vitro initiation of transcription by RNA polymerase II on in vivo-assembled chromatin templates. 131 66

The adenovirus E1A and E1B proteins are required for transformation of primary rodent cells. When expressed in the absence of the 19,000-dalton (19K) E1B protein, however, the E1A proteins are acutely cytotoxic and induce host cell chromosomal DNA fragmentation and cytolysis, analogous to cells undergoing programmed cell death (apoptosis). E1A alone can efficiently initiate the formation of foci which subsequently undergo abortive transformation whereby stimulation of cell growth is counteracted by continual cell death. Cell lines with an immortalized growth potential eventually arise with low frequency. Coexpression of the E1B 19K protein with E1A is sufficient to overcome abortive transformation to produce high-frequency transformation. Like E1A, the tumoricidal cytokine tumor necrosis factor alpha (TNF-alpha) evokes a programmed cell death response in many tumor cell lines by inducing DNA fragmentation and cytolysis. Expression of the E1B 19K protein by viral infection, by transient expression, or in transformed cells completely and specifically blocks this TNF-alpha-induced DNA fragmentation and cell death. Cosegregation of 19K protein transforming activity with protection from TNF-alpha-mediated cytolysis demonstrates that both activities are likely the consequence of the same function of the protein. Therefore, we propose that by suppressing an intrinsic cell death mechanism activated by TNF-alpha or E1A, the E1B 19K protein enhances the transforming activity of E1A and enables adenovirus to evade TNF-alpha-dependent immune surveillance.
Mol Cell Biol 1992 Jun
PMID:The 19-kilodalton adenovirus E1B transforming protein inhibits programmed cell death and prevents cytolysis by tumor necrosis factor alpha. 131 6

9-(2-Phosphonylmethoxyethyl)adenine (PMEA) is a selective and potent inhibitor of retrovirus and herpesvirus replication in vitro and in vivo. In cell culture studies, pretreatment of HeLa S3 cells with PMEA before infection enhanced its antiviral potency by almost 10-fold, compared with treatment of the cells only after viral infection. To elucidate the basis for this observation, the uptake, metabolism, and retention of PMEA metabolites were examined in uninfected and herpes simplex virus type 1-infected cells, by using [2,8-3H]PMEA. Uptake of the drug into both acid-soluble and acid-insoluble fractions was slow and did not begin to plateau until close to 24 hr. High performance liquid chromatographic analysis of acid-soluble extracts revealed at least four metabolites in addition to PMEA itself, designated as X, Y, DP, and TP. Metabolites X and Y, which were distinct from PMEA and its mono- and diphosphoryl derivatives, represented almost 90% of the radioactivity associated with the cells after 24 hr of incubation. Dephosphorylation of acid-soluble metabolites resulted in accumulation of radioactivity in the peaks associated with PMEA and X. Most of the radioactivity in the acid-insoluble fraction was associated with DNA. Enzymatic digestion of [3H] PMEA-labeled DNA from either infected or uninfected cells yielded both metabolite X and PMEA itself. The role of newly discovered PMEA metabolites in its antiviral activity and cytotoxicity is not clear.
Mol Pharmacol 1992 Sep
PMID:Cell-protecting effect against herpes simplex virus-1 and cellular metabolism of 9-(2-phosphonylmethoxyethyl)adenine in HeLa S3 cells. 132 49


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