Transcriptional regulation of Kaposi's sarcoma-associated herpesvirus-encoded oncogene viral interferon regulatory factor by a novel transcriptional silencer, Tis.
Viral interferon regulatory factor (vIRF) encoded by Kaposi's sarcoma-associated herpesvirus (KSHV) has been shown to transform NIH3T3 and Rat-1 cells, inhibit interferon signal transduction, and regulate the expression of KSHV genes. We had previously characterized the vIRF core promoter and defined a 12-O-tetradecanoylphorbol-13-acetate (TPA)-responsive region in the upstream regulatory sequence of vIRF gene. Here, we have further identified a novel transcriptional silencer, named Tis in this region. Tis represses the promoter activities of vIRF and heterologous herpes simplex virus thymidine kinase genes in both position- and orientation-independent manners. Deletion analysis has identified a cis-element of 23 nucleotides that is essential for the negative regulation. Two Tis-binding protein complexes, named vR1 and vR2, were observed by electrophoretic mobility shift assays using nuclear extracts from both KSHV-negative and -positive cell lines. A sequence fragment GAGTTAATAGGTAGAG in the cis-element was shown to be required for the DNA-protein interactions as well as the repression of vIRF promoter activity. Point-mutation analysis identified TTAAT and GTTAATAG as the core sequence motifs for the binding of vR1 and vR2, respectively. These results define the function of a novel transcriptional silencer in the regulation of vIRF gene expression.
Wang
Xin-Ping
XP
Departments of Pediatrics and Microbiology, University of Texas Health Science Center at San Antonio, San Antonio, Texas 78229-3900, USA.
Zhang
Yan-Jin
YJ
Deng
Jian-Hong
JH
Pan
Hong-Yi
HY
Zhou
Fu-Chun
FC
Gao
Shou-Jiang
SJ
D016475:3T3 Cells; D020816:Amino Acid Motifs; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D017353:Gene Deletion; D015967:Gene Expression Regulation, Viral; D020868:Gene Silencing; D017930:Genes, Reporter; D006367:HeLa Cells; D019288:Herpesvirus 8, Human / Q000378:metabolism*; D006801:Humans; D050835:Interferon Regulatory Factors; D051379:Mice; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D051381:Rats; D012097:Repressor Proteins / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014764:Viral Proteins
D004268:DNA-Binding Proteins; D050835:Interferon Regulatory Factors; D012097:Repressor Proteins; D014157:Transcription Factors; D014764:Viral Proteins; C413486:viral interferon regulatory factors; D015500:Chloramphenicol O-Acetyltransferase; D013755:Tetradecanoylphorbol Acetate