E1BF/Ku interacts physically and functionally with the core promoter binding factor CPBF and promotes the basal transcription of rat and human ribosomal RNA genes.
We have previously characterized an RNA polymerase (pol) I transcription factor, E1BF, from rat cells. This protein is immunologically related to Ku autoantigen and is required in pol-I directed transcription of rodent ribosomal RNA gene (rDNA). Glycerol density gradient fractionation and in situ UV cross-linking analysis of the purified factor showed directly that it consists of a heterodimer of 85 and 72 kDa polypeptides. E1BF also interacted with the human core promoter and augmented transcription of human rDNA as much as fivefold in HeLa nuclear extract, whereas transcription from adenovirus major late promoter, CMV or SV40 early promoters by pol II and of U6 and 5S RNA genes by pol III were either unaffected or minimally inhibited by the antibodies. Purified rat E1BF partially restored the suppression of human rDNA transcription by anti-Ku antibodies. Immunoprecipitation of rat cell extract with the anti-Ku antibodies followed by SDS-PAGE of the precipitated proteins and Southwestern analysis showed that E1BF interacts with CPBF, a core promoter binding factor. When the majority of CPBF and E1BF was removed from the reaction mixture by preincubation with a core promoter oligo nucleotide fragment, rDNA transcription was severely impaired. Addition of exogenous CPBF or E1BF to such a reaction resulted in significant restoration of the transcription, whereas inclusion of both factors caused further enhancement of rDNA transcription. These data demonstrate that E1BF is a basal pol I transcription factor that interacts with a core promoter binding factor both physically and functionally, and that is not a general pol II or pol III transcription factor.
Niu
H
H
Department of Pharmacology and Molecular Biology, Chicago Medical School, IL 60064, USA.
Zhang
J
J
Jacob
S T
ST
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D034961:Antigens, Nuclear; D004265:DNA Helicases; D004275:DNA, Ribosomal / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000494:pharmacology; D006801:Humans; D000072200:Ku Autoantigen; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D012318:RNA Polymerase I / Q000235:genetics; D012319:RNA Polymerase II / Q000235:genetics; D012320:RNA Polymerase III / Q000235:genetics; D012335:RNA, Ribosomal / Q000235:genetics*; D012341:RNA, Ribosomal, 5S / Q000235:genetics; D051381:Rats; D013045:Species Specificity; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics* / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects / Q000276:immunology
D034961:Antigens, Nuclear; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012335:RNA, Ribosomal; D012341:RNA, Ribosomal, 5S; D014157:Transcription Factors; D012319:RNA Polymerase II; D012318:RNA Polymerase I; D012320:RNA Polymerase III; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; C000606511:Xrcc6 protein, rat; D000072200:Ku Autoantigen