Transcript Cleavage in an RNA Polymerase I Elongation Complex: EVIDENCE FOR A DISSOCIABLE ACTIVITY SIMILAR TO BUT DISTINCT FROM TFIIS

Abstract

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.