Gene expression patterns determine cell identity and are therefore essential for proper development and prevention of inappropriate cell proliferation. Expression patterns are determined by transcriptional activation and repression. Mechanisms of gene expression, the focus of this research project, can be local or regional and involve alterations in chromatin structure. Locus-specific repressors affect only a few nucleosomes, whereas regional silencing involves specialized chromatin structures that propagate great distances along chromosomes. Key issues in the field are how the formation of repressive chromatin is restricted in space and time and how these chromatin structures repress transcription. To address these issues, this proposal focuses on a non-spreading locus-specific repressor protein, Sum1p from S. cerevisiae and a mutant version of this protein, Sum1-1p, which propagates along a chromosome and achieves regional silencing. Sum1p and Sum1-1p differ only a single critical amino acid and therefore present a unique opportunity to explore the similarities and differences between local and regional repression. Using a combination of biochemical and genetic approaches to investigate the mechanisms by which these almost identical proteins have such different spreading properties, we will provide fundamental insights into the regulation of spreading in regional silencing as well as shared mechanism of repression. The 4 specific aims of this research project extend a considerable body of preliminary results.
The aims are: (i) To understand how the SUM1-1 mutation changes the action of Sum1p by determing the function (s) of the conserved domain of Sum1p in which the amino acids resides, (ii) To better understand how wild-type and mutant Sum1p interact with chromatin by determining whether wild-type and/or mutant Sum1p bind to histones, (iii) To explore the role of an NAD+-dependent deacetylase in Sum1p-mediated repression, and (iv) To investigate how the spreading of Sum1-1p is regulated by studying the establishment of Sum1-1p-mediated silencing in vivo.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Molecular Genetics B Study Section (MGB)
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Carter, Anthony D
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Duke University
Schools of Medicine
United States
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Tsai, Hung-Ji; Baller, Joshua A; Liachko, Ivan et al. (2014) Origin replication complex binding, nucleosome depletion patterns, and a primary sequence motif can predict origins of replication in a genome with epigenetic centromeres. MBio 5:e01703-14
Prescott, Eugenia T; Safi, Alexias; Rusche, Laura N (2011) A region of the nucleosome required for multiple types of transcriptional silencing in Saccharomyces cerevisiae. Genetics 188:535-48
Froyd, Cara A; Rusche, Laura N (2011) The duplicated deacetylases Sir2 and Hst1 subfunctionalized by acquiring complementary inactivating mutations. Mol Cell Biol 31:3351-65
Hickman, Meleah A; Froyd, Cara A; Rusche, Laura N (2011) Reinventing heterochromatin in budding yeasts: Sir2 and the origin recognition complex take center stage. Eukaryot Cell 10:1183-92
Lynch, Patrick J; Rusche, Laura N (2010) An auxiliary silencer and a boundary element maintain high levels of silencing proteins at HMR in Saccharomyces cerevisiae. Genetics 185:113-27
Hickman, Meleah A; Rusche, Laura N (2010) Transcriptional silencing functions of the yeast protein Orc1/Sir3 subfunctionalized after gene duplication. Proc Natl Acad Sci U S A 107:19384-9
Rusche, Laura N; Rine, Jasper (2010) Switching the mechanism of mating type switching: a domesticated transposase supplants a domesticated homing endonuclease. Genes Dev 24:10-4
Rusche, Laura N; Lynch, Patrick J (2009) Assembling heterochromatin in the appropriate places: A boost is needed. J Cell Physiol 219:525-8
Hickman, Meleah A; Rusche, Laura N (2009) The Sir2-Sum1 complex represses transcription using both promoter-specific and long-range mechanisms to regulate cell identity and sexual cycle in the yeast Kluyveromyces lactis. PLoS Genet 5:e1000710
Lynch, Patrick J; Rusche, Laura N (2009) A silencer promotes the assembly of silenced chromatin independently of recruitment. Mol Cell Biol 29:43-56

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