All ETDs from UAB

Advisory Committee Chair

Igor N Chesnokov

Advisory Committee Members

David M Bedwell

Louise T Chow

Peter J Detloff

Guillermo Marques

Document Type

Dissertation

Date of Award

2010

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

DNA replication and cytokinesis are two important parts of the cell division process. Our studies address different aspects of both of these processes and potential links between them. One of the best approaches for studying DNA replication in eukaryotes has been an analysis in cell free in vitro systems. The existing cell free replication systems for Drosophila are relatively ineffective. In the first part of this work, we have developed an optimized in vitro system to study DNA replication using early egg extracts of Drosophila. Our studies indicate that this system proves to be an extremely useful tool for a functional dissection of the processes and factors involved in DNA replication in metazoans. The central component of eukaryotic DNA replication, a six-subunit origin recognition complex (ORC), is known to be involved in non-replicative functions as well. In the second part of this thesis, we analysed the interaction of Orc6, the smallest subunit of ORC, with the septin protein Pnut and the whole Drosophila septin complex. Our results suggest an active role for Orc6 in septin complex function. Orc6 might be part of a control mechanism directing the cytokinesis machinery during the final steps of mitosis. Another control mechanism of septin complex activity may be related to post-translational modifications of septins. In the final part of the thesis, we have identified and characterized a phosphorylated form of Pnut protein which can be found in the embryonic extracts in the early stages of development. We propose that phosphorylation of Pnut may inhibit activity of the septin complex to prevent cytokinesis during development of syncitial blastoderm. Together, our studies provide further characterization of key components of the replication and cytokinesis machinery, identify new regulatory mechanisms of these processes and the functional link between two major events of cell division.

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