The long-term objective of the proposed research is to understand the molecular basis for the developmental switch which controls the transition from the mitotic cell cycle to the specialized meiotic cycle. The differentiative event of meiosis is central to the process of gametogenesis and sexual reproduction, but is poorly understood. The fission yeast, Schizosaccharomyces pombe, will be used as a model organism for the study of the control of meiosis. It has a simple sexual life cycle, is amenable to biochemical analysis and has been used extensively for genetic study of mitosis and meiosis. Gene isolation by complementation of yeast genetic markers is a routine procedure in S. pombe. Two genes, ran1 and mei2, which have a central role in controlling the transition from mitosis to meiosis have previously been identified. ran1 mutants are depressed for each of the normal requirements for meiosis and sporulation, (1) heterozygosity at the mating-type locus, (2) nutritional limitation (3) diploidy. mei2 is an extragenic suppressor of ran1, which in an otherwise wildtype background, also arrests normal sexual differentiation prior to commitment to meiosis. mei2 is dominant for ran1 suppression and recessive for its defect in normal meiosis. mei2 has been isolated and ran1 will be isolated. Both genes will be sequenced to assist in (1) mapping of their transcripts, (2) insertion of a yeast selectable marker into the gene for creation of a null-mutation in yeast by the gene disruption technique, (3) computer comparison of the predicted amino acid sequence of the genes with a data bank of all known proteins. The transcripts of mei2 and ran1 will be mapped using the S1 technique and the control of their expression will be studied. In particular, the possibility that inhibition of ran1 transcription is essential for meiotic commitment will be tested. The products of the ran1 and mei2 genes will be systematically investigated by identifying the proteins by 2-D gel electrophoresis and by raising antibodies against them. Such antibodies will be used to study the levels and intracellular localization of their antigens at different stages of the life cycle and to test for modifications, such as phosphorylation. Finally, several previously isolated mutants will be screened for possible defects in cAMP-dependent protein kinase activity, since this enzyme appears to have a role in commitment to meiosis in the distantly related yeast, Saccharomyces cerevisiae.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM034517-03
Application #
3285672
Study Section
Genetics Study Section (GEN)
Project Start
1984-12-01
Project End
1987-11-30
Budget Start
1986-12-01
Budget End
1987-11-30
Support Year
3
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Cold Spring Harbor Laboratory
Department
Type
DUNS #
065968786
City
Cold Spring Harbor
State
NY
Country
United States
Zip Code
11724