This proposal focuses on chromosome synapsis and cell cycle regulation during meiosis in budding yeast. Chromosome synapsis and meiotic recombination are required for proper meiotic chromosome segregation and thus for the generation of euploid gametes. Meiotic checkpoints block nuclear division in response to defects in synapsis and recombination and thus prevent the production of aneuploid offspring. Zipi is a major structural component of the synaptonemal complex. Zip 1-interacting proteins will be identified and characterized. The significance of the DNA-binding activity of Zipi will be explored. A multi-protein ZipfMsh complex acts at sites of synapsis initiation to trigger Zipi polymerization. The structure and assembly of this complex will be investigated using the two-hybrid protein system and indirect immunofluoresence. To test the hypothesis that synapsis initiates at crossover sites, Zip/Msh complexes will be recovered by chromatin immunoprecipitation and the frequency of crossing over in the associated DNA will be measured. The role of the Zip/Msh complex in regulating crossover distribution will be investigated by (i) determining the effect of zip4 on crossover interference, and (ii) determining the effects of various zip/msh mutations on the rates of crossing over on chromosomes of different sizes. The Ssp2 and Ssp3 proteins appear to negatively regulate meiotic cell cycle progression at or prior to the initiation of recombination. ssp2 and ssp3 will be examined for their effects on (i) the lengths of meiotic S-phase and prophase, (ii) the DNA replication and homolog pairing checkpoints, and (iii) the coupling between DNA replication and recombination initiation. Ssp2 and Ssp3 will be immunolocalized. The Redi and Ddcl proteins undergo Meki-dependent phosphorylation in response to the initiation of recombination. The importance of Redi phosphorylation will be determined by identifying and mutating the phosphorylated residues. The relative importance of Redi and Ddcl phosphorylation in arresting the cell cycle at pachytene will be assessed by separating these events by mutation. Proteins that coimmunoprecipitate with Meki will be identified in order to find novel Meki substrates and regulators. The meiosis-specific Pch2 localizes to the nucleolus and is required for the pachytene checkpoint. Activity of the Cdcl4 phosphatase is regulated by sequestration in the nucleolus. The possibility that Pch2 regulates Cdcl4 activity will be tested by examining the effects of Cdcl4 inactivation and overproduction.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM028904-18
Application #
6395423
Study Section
Genetics Study Section (GEN)
Program Officer
Anderson, Richard A
Project Start
1981-04-01
Project End
2005-07-31
Budget Start
2001-08-01
Budget End
2002-07-31
Support Year
18
Fiscal Year
2001
Total Cost
$164,162
Indirect Cost
Name
Yale University
Department
Physiology
Type
Schools of Arts and Sciences
DUNS #
082359691
City
New Haven
State
CT
Country
United States
Zip Code
06520
Mitra, Neal; Roeder, G Shirleen (2007) A novel nonnull ZIP1 allele triggers meiotic arrest with synapsed chromosomes in Saccharomyces cerevisiae. Genetics 176:773-87
Li, Jing; Agarwal, Seema; Roeder, G Shirleen (2007) SSP2 and OSW1, two sporulation-specific genes involved in spore morphogenesis in Saccharomyces cerevisiae. Genetics 175:143-54
Li, Jing; Hooker, Gillian W; Roeder, G Shirleen (2006) Saccharomyces cerevisiae Mer2, Mei4 and Rec114 form a complex required for meiotic double-strand break formation. Genetics 173:1969-81
Tsubouchi, Tomomi; Zhao, Hongyu; Roeder, G Shirleen (2006) The meiosis-specific zip4 protein regulates crossover distribution by promoting synaptonemal complex formation together with zip2. Dev Cell 10:809-19
Tsubouchi, Hideo; Roeder, G Shirleen (2006) Budding yeast Hed1 down-regulates the mitotic recombination machinery when meiotic recombination is impaired. Genes Dev 20:1766-75
Tsubouchi, Tomomi; Roeder, G Shirleen (2005) A synaptonemal complex protein promotes homology-independent centromere coupling. Science 308:870-3
Hong, Eun-Jin Erica; Roeder, G Shirleen (2002) A role for Ddc1 in signaling meiotic double-strand breaks at the pachytene checkpoint. Genes Dev 16:363-76
Novak, J E; Ross-Macdonald, P B; Roeder, G S (2001) The budding yeast Msh4 protein functions in chromosome synapsis and the regulation of crossover distribution. Genetics 158:1013-25
Tung, K S; Hong, E J; Roeder, G S (2000) The pachytene checkpoint prevents accumulation and phosphorylation of the meiosis-specific transcription factor Ndt80. Proc Natl Acad Sci U S A 97:12187-92
Bailis, J M; Roeder, G S (2000) Pachytene exit controlled by reversal of Mek1-dependent phosphorylation. Cell 101:211-21

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