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Global analysis of transcriptionally engaged yeast RNA polymerase III reveals extended tRNA transcripts

ABSTRACT: RNA polymerase III (RNAPIII) synthesizes a range of highly abundant small stable RNAs, principally pre-tRNAs. Here we report the genome-wide analysis of nascent transcripts attached to RNAPIII under permissive and restrictive growth conditions. This revealed strikingly uneven polymerase distributions across transcription units, generally with a predominant 5Â´ peak. This peak was higher for more heavily transcribed genes, suggesting that initiation site clearance is rate limiting during RNAPIII transcription. Down-regulation of RNAPIII transcription under stress conditions was found to be uneven; a subset of tRNA genes showed low response to nutrient shift or loss of the major transcription regulator Maf1, suggesting potential âhousekeepingâ roles. Many tRNA genes were found to generate long, 3Â´-extended forms due to read-through of the canonical poly(U) terminators. The degree of read-through was anti-correlated with the density of T-residues in the coding strand, and multiple, functional terminators can be located far downstream. The steady-state levels of 3Â´-extended pre-tRNA transcripts are low, apparently due to targeting by the nuclear surveillance machinery; especially the RNA-binding protein Nab2, cofactors for the nuclear exosome and the 5Â´-exonuclease Rat1. CRAC protocol performed on samples containing HTP-tagged proteins: Rpc160(Rpo31), Nab2, Rrp44(Dis3), Rrp6 and Mtr4. Rpc-160-bound RNAs were analyzed in wildtype and maf1 mutant cells and after a shift to medium containing glycerol.

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Tomasz Turowski

PROVIDER: E-GEOD-77863 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Global analysis of transcriptionally engaged yeast RNA polymerase III reveals extended tRNA transcripts

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