Access Type

Open Access Dissertation

Date of Award

1-1-2010

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Pharmacology

First Advisor

George S. Brush

Abstract

Progression through meiosis occurs through a strict sequence of events, so that one round of DNA replication precedes programmed recombination and two nuclear divisions. Cyclin dependent kinase 1 (Cdk1) is required for meiosis, and any disruption in its activity leads to meiotic defects. The Cdk1 inhibitor, Sic1, regulates the G1-S transition in the mitotic cell cycle and the analogous transition in meiosis. We have employed a form of Sic1, Sic1deltaPHA, that is mutated at multiple phosphorylation sites and resistant to degradation. Meiosis specific expression of Sic1deltaPHA disrupts Cdk1 activity and leads to significant accumulation of over replicated DNA. These data suggested that Cdk1 is required to prevent inappropriate re-initiation of DNA synthesis during meiosis, as it is during mitosis. In addition, deletion of the gene DMC1, which encodes a recombinase required for meiotic recombination, prevented DNA re-replication. However, the additional deletions of RAD17, MEC1, MEK1, RED1, HOP1, or PCH2 restored the re-replication phenotype. These proteins are all individually required for the meiotic recombination checkpoint. Therefore, indicating that induction of the pachytene checkpoint by a gene deletion of DMC1 was responsible for meiotic DNA re-replication. The downstream targets of the meiotic recombination checkpoint, Ndt80, Sum1, and Swe1, which function to maintain arrest in the pachytene stage of prophase of MI, were unable to inhibit meiotic DNA re-replication induced by Sic1deltaPHA expression. Therefore, it appears that a separate branch of the pachytene checkpoint exists that has the ability to prevent extra rounds of meiotic DNA replication. We investigated whether the G1-S DNA damage checkpoint as defined in the mitotic cell cycle might be implicated, and found that this checkpoint was not involved. In summary these dissertation studies discuss the implications of lowering Cdk1 activity to induce meiotic DNA re-replication, as well as the interplay of the pachytene checkpoint. Our results provide strong evidence that the control of DNA replication is likely to be similar in mitosis and meiosis. In addition, our results are the first to show the ability of the pachytene checkpoint to monitor meiotic DNA replication.