Renaturase and ribonuclease H: a novel mechanism that influences transcript displacement by RNA polymerase II in vitro.
I have previously reported an activity in HeLa cells which facilitates transcript displacement by purified mammalian RNA polymerase II in vitro. I have shown that this activity copurifies with one of two separable ribonuclease (RNase) H activities in HeLa cells. The RNase H activity in question has characteristics similar to those reported for RNase H2b from calf thymus. RNase H proteins purified from several other sources including Escherichia coli also show renaturase activity. When the renaturase/RNase H protein is present during transcription by purified RNA polymerase II, transcripts are truncated close to the 5' end, and the remainder of the transcript is displaced normally from its template by the polymerase. Since RNA polymerase II dependent transcripts in vivo normally require the presence of the 5'-triphosphate terminus for capping, the in vivo significance of RNase H as a renaturase factor is presently not understood. However, the in vitro action of renaturase/RNase H suggests that the mechanism of this reaction may involve R-loop displacement after formation of a short single-stranded region of DNA on the template strand following hydrolysis of a hybrid transcript oligonucleotide by RNase H.
Kane
C M
CM
Department of Biochemistry, University of California, Berkeley 94720.
D000818:Animals; D002417:Cattle; D018736:DNA Adducts; D004269:DNA, Bacterial; D004330:Drosophila / Q000201:enzymology; D004722:Endoribonucleases / Q000302:isolation & purification / Q000378:metabolism*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology; D006367:HeLa Cells / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D016685:Mitomycin; D008937:Mitomycins; D008970:Molecular Weight; D012319:RNA Polymerase II / Q000378:metabolism*; D016914:Ribonuclease H; D013950:Thymus Gland / Q000201:enzymology; D014158:Transcription, Genetic
D018736:DNA Adducts; D004269:DNA, Bacterial; D008937:Mitomycins; C056417:mitomycin C-DNA adduct; D016685:Mitomycin; D012319:RNA Polymerase II; D004722:Endoribonucleases; D016914:Ribonuclease H