Identification of dTopors domains required for subcellular localization

UNCG Author/Contributor (non-UNCG co-authors, if there are any, appear on document)
Fidele Byungura (Creator)
Institution
The University of North Carolina at Greensboro (UNCG )
Web Site: http://library.uncg.edu/
Advisor
John Tomkiel

Abstract: The Topors protein is a tumor suppressor in humans that associates with and regulates a number of cell cycle regulators that include Topoisomerase I and p53. It possesses both ubiquitin and SUMO ligase activity and its mutation or downregulation has been associated with some human cancers and diseases. The Drosophila homologue, dTopors, presents structural and functional similarities to human Topors. Both proteins have four conserved domains: a RING finger domain, an RS region, PEST domains and a consensus sequence unique to Topors homologs. To gain insight into the role(s) of these domains in subcellular localization of the protein, we generated transgenic flies expressing dTopors peptides fused to Green Fluorescent Protein. These included full length dTopors protein, and truncations containing amino acids 1-967, 1-367, 1-182, 368-1038, and 968-1038. Using confocal microscopy on living cells, we localized the expressed proteins in two different cell types where dTopors function had been previously examined, salivary glands and spermatocytes. We identified and mapped two nuclear localization sequences (between aa183-367 and aa368-968), a chromosome-binding domain (aa1-367), two nuclear lamina localization domains (between aa1-367 and aa368-967), and a domain responsible for targeting dTopors to punctate nuclear spots (aa368-968). We observed some tissue-specific differences in localization patterns, suggesting that tissue-specific modifications and/or binding partners may be important for dTopors localization, and presumably, its function. These findings constitute a first step to understanding the relationship between dTopors localization and its function.

Additional Information

Publication
Thesis
Language: English
Date: 2009
Keywords
Drosophila, dTopors, Meiosis, RING finger
Subjects
Cancer $x Gene therapy $x Research.
Antineoplastic agents $x Research.
Protein binding.
DNA topoisomerase I.
Meiosis.
Drosophila.