The PCF1-1 mutation increases the activity of the transcription factor (TF) IIIB fraction from Saccharomyces cerevisiae.
PCF1-1 is a dominant suppressor of a tRNA gene A block promoter mutation (A19) in Saccharomyces cerevisiae. Transcriptional activation by PCF1-1 was examined in vitro using whole-cell extracts and purified factors derived from mutant and wild-type strains. These experiments show that PCF1 is a general activator of RNA polymerase III (pol III) gene transcription. The transcription of all pol III genes analyzed to date, including type I and numerous type II genes, is increased 3-7 fold in mutant cell extracts. Single round transcription assays indicate that the PCF1-1 mutation increases the number of functional preinitiation complexes and suggest that this is achieved by increasing the intrinsic activity of the encoded product rather than its amount. Point mutations throughout the A block of the sup3-e gene and numerous B block mutations fail to abolish transcriptional activation suggesting that interactions between TFIIIC and the internal promoter are unaffected by PCF1-1. Moreover, TFIIIC purified from the mutant strain is incapable of conferring PCF1-1 transcriptional activity to a reaction in which the remaining components are wild-type. In contrast, the activity of the TFIIIB fraction is increased in PCF1-1 extracts and can reconstitute mutant levels of transcription when added to wild-type TFIIIC and polymerase. We conclude that PCF1 is a component or regulator of TFIIIB.
Willis
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I
Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461.
Oksman
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A
López-De-León
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A
D001483:Base Sequence; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D005800:Genes, Fungal / Q000235:genetics*; D016153:Genes, Suppressor / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D012320:RNA Polymerase III / Q000235:genetics; D012343:RNA, Transfer / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D035361:Transcription Factor TFIIIB; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D019474:Transcription Factors, TFIII; D014158:Transcription, Genetic / Q000235:genetics; D000098832:Transcription Factor TFIIIC
D005656:Fungal Proteins; D035361:Transcription Factor TFIIIB; D014157:Transcription Factors; D019474:Transcription Factors, TFIII; D000098832:Transcription Factor TFIIIC; D012343:RNA, Transfer; D012320:RNA Polymerase III