Loss of a phosphorylated form of transcription factor CREB/ATF in poliovirus-infected cells.
Host cell RNA synthesis is inhibited by poliovirus infection. We have studied the mechanism of poliovirus-induced inhibition of RNA polymerase II-mediated transcription by using the adenovirus early region 3 (E3) promoter. In vitro transcription from the E3 promoter was severely inhibited in extracts prepared from poliovirus-infected HeLa cells. Four regions in the E3 promoter have been shown to serve as binding sites for cellular transcription factors. These regions contain binding sites for transcription factors NF-1 (site IV), AP-1 (site III), CREB/ATF (site II), and the TATA factor (site I). Binding to these four regions was not significantly altered by poliovirus infection as assayed by DNase I footprinting analysis; furthermore, gel retardation assays failed to reveal dramatic differences in the total amount of CREB/ATF-, AP-1-, and NF-1-binding activity present in mock- or poliovirus-infected cell extracts. Gel retardation assays, however, did reveal significant qualitative differences in the DNA-protein complexes formed with a CREB/ATF-binding site in extracts prepared from poliovirus-infected cells as compared to mock-infected cell extracts. Radioimmunoprecipitation reactions performed with antiserum against CREB/ATF revealed a severe reduction in a phosphorylated form of the protein present in poliovirus-infected cell extracts. However, in vitro kinase reactions demonstrated that mock- and poliovirus-infected cell extracts contained similar levels of CREB/ATF. Expression from the E3 promoter was shown to be activated by CREB/ATF in vivo; this induction was dependent upon the phosphorylation of CREB/ATF. Thus, we propose that poliovirus infection inhibits transcription from the E3 promoter, at least in part, through the dephosphorylation of CREB/ATF.
Kliewer
S
S
Department of Microbiology and Immunology, University of California, Los Angeles 90024.
Muchardt
C
C
Gaynor
R
R
Dasgupta
A
A
D051696:Activating Transcription Factors; D001483:Base Sequence; D001798:Blood Proteins / Q000235:genetics* / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002472:Cell Transformation, Viral; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D003850:Deoxyribonuclease I; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics*; D015345:Oligonucleotide Probes; D010710:Phosphates / Q000378:metabolism; D010766:Phosphorylation; D017955:Poliovirus / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011494:Protein Kinases / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
D051696:Activating Transcription Factors; D001798:Blood Proteins; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D015345:Oligonucleotide Probes; D010710:Phosphates; D014157:Transcription Factors; D011494:Protein Kinases; D003850:Deoxyribonuclease I