title: Point mutations in the Rpb9-homologous domain of Rpc11 that impair transcription termination by RNA polymerase III.
abstract: RNA polymerase III recognizes and pauses at its terminator, an oligo(dT) tract in non-template DNA, terminates 3' oligo(rU) synthesis within this sequence, and releases the RNA. The pol III subunit Rpc11p (C11) mediates RNA 3'-5' cleavage in the catalytic center of pol III during pausing. The amino and carboxyl regions of C11 are homologous to domains of the pol II subunit Rpb9p, and the pol II elongation and RNA cleavage factor, TFIIS, respectively. We isolated C11 mutants from Schizosaccharomyces pombe that cause pol III to readthrough terminators in vivo. Mutant RNA confirmed the presence of terminator readthrough transcripts. A predominant mutation site, F32, resides in the C11 Rpb9-like domain. Another mutagenic approach confirmed the F32 mutation and also isolated I34 and Y30 mutants. Modeling Y30, F32 and I34 of C11 in available cryoEM pol III structures predicts a hydrophobic patch that may interface with C53/37. Another termination mutant, Rpc2-T455I, appears to reside internally, near the RNA-DNA hybrid. We show that the Rpb9 and TFIIS homologous mutants of C11 reflect distinct activities, that differentially affect terminator recognition and RNA 3' cleavage. We propose that these C11 domains integrate action at the upper jaw and center of pol III during termination.
authors: Intramural Research Program, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Commissioned Corps, US Public Health Service, Bethesda, MD, USA. 20892
authors: James R
authors: JR
authors: Iben
authors: Julie K
authors: JK
authors: Mazeika
authors: Sam
authors: S
authors: Hasson
authors: Keshab
authors: K
authors: Rijal
authors: Aneeshkumar G
authors: AG
authors: Arimbasseri
authors: Amy N
authors: AN
authors: Russo
authors: Richard J
authors: RJ
authors: Maraia
mesh_terms: D000595:Amino Acid Sequence; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010641:Phenotype; D017354:Point Mutation; D011071:Poly T / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry; D012320:RNA Polymerase III / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012343:RNA, Transfer / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry; D012568:Schizosaccharomyces / Q000235:genetics; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D013728:Terminator Regions, Genetic; D014158:Transcription, Genetic
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D011071:Poly T; D012343:RNA, Transfer; D012319:RNA Polymerase II; C509257:Rpb9 protein, S cerevisiae; C116192:Rpb9 protein, S pombe; D012320:RNA Polymerase III