HYDROLYTIC CLEAVAGE OF NASCENT RNA IN RNA POLYMERASE-III TERNARY TRANSCRIPTION COMPLEXES

Highly purified yeast RNA polymerase III ternary complexes were found to possess a hydrolytic chain retracting activity that cleaves nascent RNA from its 3'-OH end. Most of the shortened transcripts were capabl e of resuming RNA chain elongation, indicating that they remain stably associated with the enzyme-DNA complex. Analysis of the products of c leavage indicated that retraction primarily occurred in dinucleotide i ncrements, but that mononucleotides were also excised at lower frequen cy. The ribonuclease activity was totally dependent on the presence of a divalent cation and was stimulated by the addition of non-cognate r ibonucleotides. The inclusion of ATP in the reaction enhanced both the rate and extent of transcript cleavage. Evidence suggesting that the hydrolytic activity is intrinsic to RNA polymerase III and factor-inde pendent is also presented. Transcript cleavage by RNA polymerase III t ernary complexes appears to be more closely related to the intrinsic n ucleolytic activity of vaccinia virus RNA polymerase ternary complexes than to TFIIS-dependent cleavage that has been described for RNA poly merase II ternary complexes.