We have been studying a class of temperature-sensitive (ts) embryonic lethal mutants from C. elegans that arrest in metaphase of meiosis I. In wildtype animals, oocytes in prophase of meiosis I are fertilized by sperm. Following fertilization, the oocyte chromosomes undergo two meiotic divisions, discarding the extra chromosomes in the polar bodies. These first meiotic divisions are important as any errors in chromosome segregation at this stage can lead to embryos with an abnormal number of chromosomes, which would likely lead to lethality. In our mutants, the oocyte chromosomes arrest in metaphase of meiosis I and never separate and never extrude polar bodies. In order to molecularly identify the genes required for the first meiotic division, we have mapped our mutants and sequenced candidate genes. In collaboration with Dr. Diane Shakes (College of William and Mary, Williamsburg, VA) and Dr. Jill Schumacher (University of Houston), we have determined that 4 of the 6 genes encode subunits of the Anaphase Promoting Complex (APC). This complex serves as an E3 ubiquitin ligase that target proteins for destruction during the metaphase to anaphase transition of the cell cycle. We have named our mutants """"""""mat"""""""" for their defects in the metaphase to anaphase transition during meiosis I. These ts mutants also display defects in spermatocyte meiosis; primary spermatocytes arrest in metaphase of meiosis I with a normal meiotic spindle, yet fail to separate chromosome homologs. Thus, these mutants disrupt meiosis in both oocytes and spermatocytes. To address the role of the mat genes in mitosis, we have examined germline proliferation and have observed cell cycle delays. Germ cells appear to spend a greater time in M-phase. We believe this delay is due to the cells having difficulty progressing through anaphase, and not due to an absolute arrest. Temperature shift-up experiments during embryogenesis do not result in embryonic phenotypes, however, somatic defects in the gonad, vulva, and male tail are apparent. This observation further suggests that mitotic divisions in the soma are affected by the mat mutants. For many of the alleles, these shift-up experiments also result in sterility. To further understand the role of these genes during development, we are characterizing their expression patterns. Genetic suppression screens are also planned in order to identify regulators or substrates of these APC subunits.
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