Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To determine the mechanisms involved in the regulation of human cytomegalovirus early gene expression, we have examined the gene that encodes the viral DNA polymerase (UL54, pol). Our previous studies demonstrated that sequences required for activation of the pol promoter by immediate-early proteins are contained within a region from -128 to +20 and that cellular proteins can bind to this activation domain. In this study, we demonstrate by competition analysis that binding of cellular proteins to pol is associated with an 18-bp region containing a single copy of a novel inverted repeat, IR1. Time course analysis indicated that viral infection increased the level of protein binding to IR1, concurrent with the activation of the pol promoter. Mutation of the IR1 element abrogated binding of cellular factors to the pol promoter and reduced by threefold the activation by immediate-early proteins. Similarly, mutation of IR1 rendered the promoter poorly responsive to activation by viral infection. Mutation of additional sequence elements in the pol promoter had little effect, indicating that IR1 plays the major role in pol promoter regulation. These studies demonstrate that the interaction between cellular factors and IR1 is important for the regulation of expression of the polymerase gene by viral proteins.
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PMID:Identification of sequence elements in the human cytomegalovirus DNA polymerase gene promoter required for activation by viral gene products. 820 92

Previous analysis of the human cytomegalovirus (HCMV) DNA polymerase (UL54) early gene promoter demonstrated that transcriptional activation of this gene is dependent upon the interaction of cellular transcription factors with viral transactivators (J. A. Kerry, M. A. Priddy, T. Y. Jervey, C. P. Kohler, T. L. Staley, C. D. Vanson, T. R. Jones, A. C. Iskenderian, D. G. Anders, and R. M. Stenberg, J. Virol. 70:373-382, 1996). A sequence element, IR1, was shown to be the primary regulatory element of this promoter in transient assays. However, assessment of this element in the context of the viral genome revealed IR1-independent activation at late times after infection. To extend these studies, we aim to identify additional sequence elements involved in the activation of the UL54 promoter. Our present studies demonstrate that the level of binding of proteins to the ATF site in the UL54 promoter is enhanced by viral infection. Furthermore this increase is sensitive to treatment with phosphonoacetic acid (PAA), a DNA synthesis inhibitor. These data suggest that the increase in the level of ATF binding activity is regulated, either directly or indirectly, by HCMV late gene expression. By using specific antibodies, we determined that ATF-1 was a major component of the proteins binding to the UL54 ATF site at late times. In addition, we have demonstrated direct binding of recombinant ATF-1 to the UL54 ATF site. To assess the biological significance of these events, a recombinant virus construct was generated that contained the UL54 promoter with a mutation in the ATF site regulating expression of the chloramphenicol acetyltransferase (CAT) reporter gene inserted between open reading frames US9 and US10. Analysis of this virus (RVATFmCAT) revealed that mutation of the ATF site does not alter the kinetics of UL54 promoter activation. However, levels of CAT mRNA and activity were reduced by 5- to 10-fold compared to those of the wild-type promoter at all stages of infection. These findings indicate that ATF-1 can regulate the levels of UL54 promoter activity at both early and late times. Furthermore, these results imply that HCMV can regulate the activity of cellular factors involved in early gene regulation.
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PMID:The role of ATF in regulating the human cytomegalovirus DNA polymerase (UL54) promoter during viral infection. 903 45

Human cytomegalovirus (HCMV) gene expression is highly cell and tissue specific. Cell factor-mediated regulatory interactions are involved in regulating the restricted expression of the HCMV major immediate-early (IE) gene (J. F. Baskar, P. P. Smith, G. Nilaver, R. A. Jupp, S. Hoffmann, N. J. Peffer, D. J. Tenney, A. M. Colberg-Poley, P. Ghazal, and J. A. Nelson, 70:3207-3213, 1996). To gain an understanding of HCMV early gene activation, we studied the effect of each of the three major IE proteins, IE72, IE86, and IE55, on the HCMV DNA polymerase gene (pol; UL54) promoter. In transient-expression assays, the IE86 protein alone was able to transactivate the pol promoter, but IE72 and IE55 were not, in permissive U373MG cells. However, we were unable to detect IE86-mediated transactivation in nonpermissive HeLa or C33-A cells. Using electrophoretic mobility shift assays (EMSAs), we found that expression of the IE86 protein in U373MG cells resulted in specific binding of a DNA complex to an inverted-repeat element, IR1, of the pol promoter. Antibody supershifting and EMSA-Western blotting experiments further showed that IE86 and the cellular transcription factor Sp1 were components of the IR1 DNA-binding complex. Furthermore, we found that binding of DNA by Sp1 was dramatically increased in the presence of IE86. Interestingly, this IE86-induced DNA-binding activity of Sp1 was inhibited by a repressor activity presented in HeLa cells. In summary, our study suggests that a viral regulatory protein can modulate the DNA binding activity of a cellular transcription factor, resulting in cell-specific transactivation of viral genes.
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PMID:Transcription factor Sp1 mediates cell-specific trans-activation of the human cytomegalovirus DNA polymerase gene promoter by immediate-early protein IE86 in glioblastoma U373MG cells. 942 Feb 20