In the yeast, Saccharomyces cerevisiae, the silent mating loci, HMLalpha and HMRa, are permanently inactivated in normal cells. This repression is part of the mechanism which generates different cell types (a, alpha, diploid) and allows mating to occur. Silencing occurs between specific DNA boundaries, termed 'silencers', and requires the function of at least the four proteins Sir1, Sir2, Sir3 and Sir4. Recently, we found that deletions in the extremely conserved histone H4 N-terminus of yeast (but not similar deletions in the N-termini of histones H2A or H2B) activate the silent mating loci specifically. We propose here experiments which will help uncover many of the details in the mechanism of chromosomal repression of the silent mating loci. Using directed mutagenesis we intend to determine a) which sequences in histone H4 are required for silencing and whether there is special significance to H4 N-terminal acetylation in modifying repression of the silent mating loci. b) We will determine whether histones are involved in other mating locus specific functions such as gene conversion, autonomous replication and segregation. c) Antibodies to Sir proteins will be used to determine whether these repressor proteins interact directly with silencer chromatin isolated as episomes and in particular whether they interact with the H4 N-terminus. d) We will ask whether there are other, as yet undefined factors which interact with H4 to repress the silent mating loci. This will be done by the identification of genes whose products suppress histone gene mutations. e) We will determine whether it is H4 alone or the H3- H4 complex which is involved in repression of HMLalpha and HMRa. The chromosomal repression of the silent mating loci in yeast has analogies to the inactivation of single copy genes seen in heterochromatin of flies and mammals. This too is accompanied by changes in chromatin structure and results in long-term genetic inactivation. The understanding of the molecular basis of silencing, is likely to have a profound effect on our understanding of terminal repression, and therefore cellular differentiation, in all eukaryotes. This knowledge should impact our ability to deal with major illness, such as cancer, in which cellular differentiation is abnormal.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM042421-05
Application #
2181363
Study Section
Genetics Study Section (GEN)
Project Start
1989-07-01
Project End
1995-03-31
Budget Start
1993-07-01
Budget End
1995-03-31
Support Year
5
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of California Los Angeles
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
119132785
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Xu, Heng-hao; Su, Trent; Xue, Yong (2016) Histone H3 N-terminal acetylation sites especially K14 are important for rDNA silencing and aging. Sci Rep 6:21900
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Xu, Feng; Zhang, Qiongyi; Zhang, Kangling et al. (2007) Sir2 deacetylates histone H3 lysine 56 to regulate telomeric heterochromatin structure in yeast. Mol Cell 27:890-900
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Xu, Feng; Zhang, Kangling; Grunstein, Michael (2005) Acetylation in histone H3 globular domain regulates gene expression in yeast. Cell 121:375-85
Kurdistani, Siavash K; Grunstein, Michael (2003) In vivo protein-protein and protein-DNA crosslinking for genomewide binding microarray. Methods 31:90-5
Carmen, Andrew A; Milne, Lisa; Grunstein, Michael (2002) Acetylation of the yeast histone H4 N terminus regulates its binding to heterochromatin protein SIR3. J Biol Chem 277:4778-81

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