Identification of a mammalian RNA polymerase I holoenzyme containing components of the DNA repair/replication system.
Traditional models for transcription initiation by RNA polymerase I include a stepwise assembly of basic transcription factors/regulatory proteins on the core promoter to form a preinitiation complex. In contrast, we have identified a preassembled RNA polymerase I (RPI) complex that contains all the factors necessary and sufficient to initiate transcription from the rDNA promoter in vitro. The purified RPI holoenzyme contains the RPI homolog of TFIID, SL-1 and the rDNA transcription terminator factor (TTF-1), but lacks UBF, an activator of rDNA transcription. Certain components of the DNA repair/replication system, including Ku70/80, DNA topoisomerase I and PCNA, are also associated with the RPI complex. We have found that the holo-enzyme supported specific transcription and that specific transcription was stimulated by the RPI transcription activator UBF. These results support the hypothesis that a fraction of the RPI exists as a preassembled, transcriptionally competent complex that is readily recruited to the rDNA promoter, i.e. as a holoenzyme, and provide important new insights into the mechanisms governing initiation by RPI.
Hannan
R D
RD
Cancer Research Centre and Department of Biochemistry, Laval University, Hotel-Dieu de Quebec, 11 Cote du Palais, Quebec G1R 2J6, Canada.
Cavanaugh
A
A
Hempel
W M
WM
Moss
T
T
Rothblum
L
L
D000818:Animals; D034961:Antigens, Nuclear; D004265:DNA Helicases; D004260:DNA Repair / Q000235:genetics; D004261:DNA Replication / Q000235:genetics; D004264:DNA Topoisomerases, Type I / Q000302:isolation & purification / Q000378:metabolism; D004275:DNA, Ribosomal / Q000235:genetics; D004268:DNA-Binding Proteins / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D020035:Holoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D000072200:Ku Autoantigen; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism; D035463:Pol1 Transcription Initiation Complex Proteins; D018809:Proliferating Cell Nuclear Antigen / Q000302:isolation & purification / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D012318:RNA Polymerase I / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014157:Transcription Factors / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics; D014407:Tumor Cells, Cultured
D034961:Antigens, Nuclear; D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D020035:Holoenzymes; D009097:Multienzyme Complexes; D009687:Nuclear Proteins; D035463:Pol1 Transcription Initiation Complex Proteins; D018809:Proliferating Cell Nuclear Antigen; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C063844:transcription factor UBF; C063660:transcription initiation factor TIF-IB; D012318:RNA Polymerase I; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; C000606511:Xrcc6 protein, rat; D000072200:Ku Autoantigen; D004264:DNA Topoisomerases, Type I