Identification and Characterization of Components of the Mitotic Spindle Checkpoint Pathway in Fission Yeast
Abstract
During anaphase of mitosis, sister chromatids are separated by the dynamic mitotic spindle. Failure to equally distribute the chromosomal DNA can lead to genetic loss and aneuploidy. In eukaryotic cells, the spindle assembly checkpoint monitors the attachments between chromatids and the spindle to initiate a cell cycle delay if the attachments are defective. The fission yeast, Schizosaccharomyces pombe, is a useful system for discovering and characterizing components of this regulatory pathway because genetic approaches can be coupled with excellent cytology. Previous work in the Sazer laboratory identified the first two spindle checkpoint genes in S. pombe: mad2 and mph1. Cells in which the mphl or mad2 gene is deleted are sensitive to the microtubule inhibitor, thiabendazole (TBZ), because they fail to arrest at metaphase to repair the spindle, and therefore undergo a lethal division. Overexpression of either mad2 or mphl activates the spindle checkpoint, arrests cells prior to anaphase, and is toxic to wild type cells. mphl acts upstream of mad2, and the toxic effect of mphl overexpression is suppressed by a mad2 deletion. This project aims to discover and characterize novel spindle checkpoint genes as suppressors of mphl overexpression. The mph1 overexpression screen has yielded mutations in three known spindle checkpoint genes and a mutation, called 4101, in an apparently novel component of the spindle checkpoint pathway. 4101 mutant cells are checkpoint deficient as indicated by their failure to arrest the cell cycle in response to spindle disruption. 4101 cells also exhibit temperature sensitive lethality, and mitotic defects are observed at the restrictive temperature in the absence of TBZ. Plasmid rescue of the temperature sensitive lethality will be exploited to identify the gene mutated in strain 4101.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jul 01, 2001
- Accession Number
- ADA396484
Entities
People
- Shelia C. Kadura
- Shelly Sazer
Organizations
- Baylor College of Medicine