Transcript cleavage in an RNA polymerase I elongation complex. Evidence for a dissociable activity similar to but distinct from TFIIS.
Stalled Xenopus RNA polymerase I (pol I) elongation complexes bearing a 52-nucleotide RNA were prepared by promoter-initiated transcription in the absence of UTP. When such complexes were isolated and incubated in the presence of Mg2+, the associated RNA was shortened from the 3'-end, and mono- and dinucleotides were released. Shortened transcripts were still associated with the DNA and were quantitatively reelongated upon addition of NTPs. The cleavage activity could be removed from the pol I-ternary complex with buffers containing 0.25% Sarkosyl. These findings indicate that a factor with characteristics similar to elongation factor TFIIS is associated with the pol I elongation complex. However, addition of recombinant Xenopus TFIIS to Sarkosyl-washed pol I elongation complexes had no effect, whereas it showed the expected effects in control reactions with identically prepared pol II elongation complexes. The results thus suggest the existence of a pol I-specific cleavage/elongation factor. I also report the sequence of a novel type of Xenopus TFIIS. The predicted amino acid sequences of the present and previously identified Xenopus TFIIS are less than 65% conserved. Thus, like mammalian species, Xenopus has at least two highly divergent forms of TFIIS.
Labhart
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Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, California 92037, USA. plabhart@scripps.edu
D000595:Amino Acid Sequence; D000818:Animals; D004257:DNA Polymerase II / Q000378:metabolism; D003902:Detergents / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000378:metabolism*; D010441:Peptide Chain Elongation, Translational; D012318:RNA Polymerase I / Q000378:metabolism*; D012521:Sarcosine / Q000031:analogs & derivatives / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D035165:Transcription Factors, General; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors; D014981:Xenopus
D003902:Detergents; D009363:Neoplasm Proteins; D014157:Transcription Factors; D035165:Transcription Factors, General; D035602:Transcriptional Elongation Factors; C018648:transcription factor S-II; C025231:sarkosyl; D012318:RNA Polymerase I; D004257:DNA Polymerase II; D012521:Sarcosine