Contractions of the C-Terminal Domain of Saccharomyces cerevisiae Rpb1p Are Mediated by Rad5p

Abstract The C-terminal domain (CTD) is an essential domain of the largest subunit of RNA polymerase II, Rpb1p, and is composed of 26 tandem repeats of a seven-amino acid sequence, YSPTSPS. Despite being an essential domain within an essential gene, we have previously demonstrated that the CTD codin...

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Veröffentlicht in:G3 : genes - genomes - genetics 2020-07, Vol.10 (7), p.2543-2551
Hauptverfasser: Stewart, Taylor, Exner, Alexandra E, Patnaik, Paras, Fuchs, Stephen M
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Sprache:eng
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Zusammenfassung:Abstract The C-terminal domain (CTD) is an essential domain of the largest subunit of RNA polymerase II, Rpb1p, and is composed of 26 tandem repeats of a seven-amino acid sequence, YSPTSPS. Despite being an essential domain within an essential gene, we have previously demonstrated that the CTD coding region is genetically unstable. Furthermore, yeast with a truncated or mutated CTD sequence are capable of promoting spontaneous genetic expansion or contraction of this coding region to improve fitness. We investigated the mechanism by which the CTD contracts using a tet-off reporter system for RPB1 to monitor genetic instability within the CTD coding region. We report that contractions require the post-replication repair factor Rad5p but, unlike expansions, not the homologous recombination factors Rad51p and Rad52p. Sequence analysis of contraction events reveals that deleted regions are flanked by microhomologies. We also find that G-quadruplex forming sequences predicted by the QGRS Mapper are enriched on the noncoding strand of the CTD compared to the body of RPB1. Formation of G-quadruplexes in the CTD coding region could block the replication fork, necessitating post-replication repair. We propose that contractions of the CTD result when microhomologies misalign during Rad5p-dependent template switching via fork reversal.
ISSN:2160-1836
2160-1836
DOI:10.1534/g3.120.401409