The PKR kinase promoter binds both Sp1 and Sp3, but only Sp3 functions as part of the interferon-inducible complex with ISGF-3 proteins.
The protein kinase regulated by RNA (PKR) is an important mediator of the antiviral and antiproliferative actions of interferon (IFN). The promoter of the PKR gene contains a novel 15-bp element designated KCS that is required for both basal and IFN-inducible transcription, with KCS function dependent upon both position and orientation relative to the ISRE element. Novel inducible protein complexes (iKIBP1, iKIBP2) that require both the KCS and the ISRE element sequences for their formation have been identified and characterized. Transcription factors Sp1 and Sp3 were found to be KCS-binding proteins by electrophoretic mobility shift analyses (EMSA) and Sepharose bead-KCS oligonucleotide pull-down assays. However, only Sp3 but not Sp1 was a constituent of the inducible iKIBP complexes. EMSA also identified STAT1, STAT2, and IRF-9 as components of the iKIBP complexes, indicating that ISGF-3 participates in iKIBP complex formation. Proteins bound at the KCS element in the absence of ISRE were able to recruit both STAT1 and STAT2 to the KCS element; recruitment was dependent upon IFN-alpha treatment. Chromatin immunoprecipitation assays revealed that the binding of Sp3, similar to STAT1 and STAT2, at the PKR promoter in vivo was IFN-dependent, but that Sp1 binding was not dependent upon IFN treatment. These results, taken together, strongly suggest a role for Sp1 in basal and Sp3 in inducible transcription of PKR and that a potential function of the KCS element is to facilitate the recruitment of ISGF-3 complex components to the PKR promoter to stimulate transcription.
Ward
Simone Visosky
SV
Department of Molecular, Cellular, and Developmental Biology, University of California, Santa Barbara 93106, USA.
Samuel
Charles E
CE
D002460:Cell Line; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D050832:Interferon-Stimulated Gene Factor 3; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D007372:Interferons / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D050794:STAT1 Transcription Factor; D050795:STAT2 Transcription Factor; D015398:Signal Transduction; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D051705:Sp3 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; C494227:IRF9 protein, human; D050832:Interferon-Stimulated Gene Factor 3; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D016898:Interferon-alpha; C497121:SP3 protein, human; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050795:STAT2 Transcription Factor; C494084:STAT2 protein, human; D016329:Sp1 Transcription Factor; D015534:Trans-Activators; D014157:Transcription Factors; D051705:Sp3 Transcription Factor; D007371:Interferon-gamma; D007372:Interferons; D019892:eIF-2 Kinase