The capped U6 small nuclear RNA is transcribed by RNA polymerase III

Abstract

U6 RNA is an abundant, capped, small nuclear RNA (snRNA) species associated with heterogeneous nuclear ribonucleoproteins in eukaryotic cells. U4 RNA and U6 RNA are hydrogen bonded in a 1:1 ratio in discrete small nuclear ribonucleoprotein particles that are required in pre-mRNA processing. Previous reports have established that the mRNAs and U1 to U5 U-snRNAs are synthesized by RNA polymerase II. Evidence is presented here for synthesis of U6 RNA by RNA polymerase III. The synthesis of U6 RNA in vitro, using Novikoff hepatoma or HeLa whole cell extracts, was not inhibited at low (1 microgram/ml) concentrations of alpha-amanitin, and only 35% inhibition occurred at 10 micrograms/ml concentration. The in vitro synthesized U6 RNA, like other RNA polymerase III transcripts, was associated with La antigen. The U6 RNA synthesized in vitro by the whole cell extracts was capped, but no other internal post-transcriptional modifications were found. Uridylic acid residues were also added post-transcriptionally to the 3'-end of U6 RNA in vitro. U6 RNA, though capped on its 5'-end, is transcribed by RNA polymerase III; this is the first report of a capped RNA molecule synthesized by RNA polymerase III.