The Role of PIF1 Helicase in Minisatellite Variation.
Reinfeld, Bradley I.
- The largest subunit of the yeast RNA polymerase II complex, RPO21, contains a tandemly repeated heptapeptide sequence whose posttranslational modifications are key to properly orchestrate transcription. Sequencing of the RPO21 locus from 36 wild strains of budding yeast uncovered many polymorphisms that resulted in different lengths of its essential C-terminal domain (CTD) repeat. An alignment of ... read morethe newly found variants demonstrated that over evolutionary time, there have been many expansions and contractions to the CTD resulting in the variety of alleles seen today. With this surprising finding of variability in an essential domain, it became important to find factors that could contribute to this observed variation. From analyzing the nucleotide sequence of RPO21, it was found that a G-quadruplex could form at the most C-terminal repeats of the CTD. Additionally this nucleotide region of the CTD has been found to be bound b ythe G4 speicifc helicas PIF1 by previous chromatin immunoprecipitation studies,. Using the tTa-dependent expression system developed in the Fuchs Lab with a pif1-m2 mutant in a Luria-Delbrück fluctuation assay, it was found that PIF1 acts an inhibitor of expansion events in the CTD. This finding led to a larger survey of the yeast genome for other minisatellites that may behave like the CTD due to the presence of PIF1 binding as well as the possibility to form a G4. This analysis ultimately found that three key transcriptional enzymes SPT5, GAL11, and RPO21 all have G4 sequences in their variable minisatellites. However, future examination is required to understand why these three repeats demonstrate polymorphisms while other similar repeats do not.read less