DNA is replicated during S phase of the S. pombe cell cycle so that each chromosome is composed of two identical sister chromatids. During M phase condensed chromosomes align at the cells equator and sister chromatids are attached to microtubules emanating from opposite spindle poles. At anaphase, sister chromatids are triggered to separate rapidly and completely and move to opposite ends of the dividing cell in the equational pattern of nuclear division forming two identical daughters. During sexual development pre-meiotic replication is followed by homologous recombination and two successive rounds of nuclear division, MI and MIT. During MI sister chromatids remain joined and homologues migrate to opposite spindle poles (reductional division). During MIl, sister chromatids separate as in mitosis, forming four haploid spores enclosed in an ascus. Cohesin is a multi-subunit complex that controls chromosome segregation by providing a physical linkage between sister chromatids. Cohesin is loaded onto newly replicated chromosomes during G liS phase and associates with chromatin both at the centromeres and chromosome arms. When cohesion is dissolved at the onset of anaphase (meiotic anaphase II), sister chromatid separation is triggered. In S. pombe Rad21p is an essential cohesion protein first identified for its role DNA repair, while Rec8p cohesin is essential for meiosis, having additional roles in homologous recombination and spore formation. Both Rad21p and Rec8p occupy critical positions in the mitotic and meiotic cohesin complexes, respectively. The aim of this study was to investigate the mitotic and meiotic functions of Rad21p and Rec8p. To this end their specific patterns of expression were exchanged so that rad21+ was expressed during meiotic development and rec8+ during the mitotic cell cycle. This was achieved by swapping their promoter regions so that each would be ectopically expressed both at physiological levels and at the appropriate time in the cell cycle. Replacing Rad21p with Rec8p during the mitotic cell cycle was found to rescue the lethal effect of a rad21+ deletion with cells having wild type morphology. No S phase delay was detected in these cells though a ~5% increased division time was seen. This rescue was also accompanied with a ~30% loss of viability, and a proportion of cells exhibited the lethal rad21 null phenotype. 111 In the absence of Rec8p cells underwent aberrant meioses with abnormal chromosome segregation, reduced spore viability and impaired ascospore development. When Rec8p was replaced with Rad21p, although aberrant meiotic divisions occurred, 66% of asci contained four DNA nuclei in comparison to 51 % for rec8 null meioses. Furthermore, ascospores developed normally and spore viability was improved by ~30% when compared to rec8 null. No delay in S phase progression was detected in cells lacking Rec8p, while expressing Rad21 p in place of Rec8p also did not alter S phase timing. These data demonstrate that the essential mitotic function of Rad21 p can be partially complemented by the meiotic cohesin Rec8p. Conversely, the role ofRec8p in ascospore development appeared to be fully complemented by the mitotic cohesin Rad21 p. Taken together, the findings of this study indicate that, while S. pombe Rad21p and Rec8p cohesins have specialised mitotic and meiotic roles, the functional overlap between them is greater than previously described. They also suggest a novel role for Rad21 p in septation and/ or ascospore development.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:398642 |
| Date | January 2003 |
| Creators | Riddell, Tracy |
| Publisher | University of Glasgow |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://theses.gla.ac.uk/5428/ |
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