Sarah Tripplehorn - Arndt and Hainer Lab
Investigating co-transcriptional regulation of the nucleosome remodeler Chd1
Nucleosomes impede DNA-templated processes and enforce transcriptional fidelity by preventing transcription from aberrant start sites. The nucleosome remodeler Chd1 dynamically controls nucleosome positioning genome-wide, but it is unclear how Chd1 function is regulated and whether Chd1 recruitment is related to the transcription cycle. Previously, our lab found that the transcription elongation factor Rtf1, a member of the Paf1 Complex (Paf1C), is in part responsible for Chd1 occupancy on chromatin in Saccharomyces cerevisiae. Rtf1 is also required for the co-transcriptional deposition of histone modifications, and contributes to Paf1C recruitment to RNA Polymerase II transcription elongation machinery. Through site-directed mutagenesis and yeast two-hybrid experiments, I have localized the interacting regions of Rtf1 and Chd1 and designed point mutations in Rtf1 that disrupt the interaction, while maintaining other functions of Rtf1. Through chromatin immunoprecipitation experiments, I found that the C-terminus of Chd1, an understudied domain, and the N-terminal region of Rtf1 are required for the Rtf1-Chd1 interaction and for proper occupancy of Chd1 on chromatin. Point mutations in RTF1 that disrupt the interaction with Chd1 lead to cryptic intragenic transcription, demonstrating the importance of this interaction for proper transcriptional control. Focusing on the N-terminal region of Rtf1, forthcoming work will address the consequences of separation-of-function mutations in RTF1 on global nucleosome positioning and Chd1 occupancy genome-wide and test the evolutionary conservation of the Rtf1 and Chd1 roles in mammalian embryonic stem cells.
Friday, December 10th, 2021
A219B Langley Hall
12:00 PM