Tuesday, January 14, 2025
1/14/2025
Mechanisms of Heterochromatic Transcriptional Gene Silencing Project Summary / Abstract Heterochromatin plays a crucial role in the repression of transposons, genome stability, and cell differentiation through its ability to silence DNA transcription. How heterochromatin represses transcription by RNA polymerase II (Pol II) is an open question. In Schizosaccharomyces pombe, histone three lysine 9 di- and tri-methyl (H3K9me) bearing heterochromatic loci accomplish transcriptional silencing by recruiting the Heterochromatin Protein 1 (HP1) homologs Swi6 and Chp2. Together, HP1swi6/chp2 proteins induce chromatin compaction, recruit downstream factors such as histone deacetylases (HDAC) and chromatin remodelers, and promote nascent RNA decay. Previous studies have unraveled the mechanisms responsible for processing and degrading nascent RNA transcripts. However, the mechanism(s) by which heterochromatin inhibits Pol II activity, referred to as transcriptional gene silencing (TGS), is not fully understood. In the proposed work, I explore the following questions: Which HP1swi6/chp2 functions are sufficient for TGS and what is the mechanism of HP1swi6/chp2 mediated TGS? I hypothesize that HP1swi6/chp2 mediates TGS through distinct activities: intrinsic nonspecific DNA binding, liquid-liquid phase separation formation, and the downstream recruitment of histone deacetylase and remodeling activities. To test this hypothesis, I will 1) identify which Swi6 functions are necessary for heterochromatic TGS using HP1swi6/chp2 mutant strains, 2) identify activities downstream of HP1swi6/chp2 that are sufficient for heterochromatic TGS by direct recruitment of proteins to H3K9me loci in cells lacking recruitment of downstream factors, and 3) Identify the specific transcriptional stage inhibited by HP1swi6/chp2 by characterizing the proteome of heterochromatic and euchromatic ectopic locus. Achieving these aims would contribute to understanding the mechanisms of gene regulation and transposon repression. In addition, mechanistic knowledge of heterochromatic TGS could help treat diseases associated with heterochromatin defects.
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