The discovery of a novel 105kDa complex comprised of BUB3, C-JUN and SUMO-1 and its relationship to ovarian cancer development

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Abstract (if available)

Abstract This work is based on the hypothesis that post-translational modifications catalyzed by ubiquitin and SUMO play an important role in tumor development and progression. Nuclear protein extracts were obtained from a cell strain transfected with SV40 large T antigen derived from a benign epithelial ovarian tumor (ovarian cystadenoma) and from a subclone of the same strain that had undergone spontaneous immortalization in vitro. Both nuclear extracts were analyzed by two-dimensional gel electrophoresis followed by western blotting with antibodies against ubiquitin and SUMO-1. Two protein complexes, respectively 40kDa and 105kDa, appeared to be differentially ubiquitinated and sumoylated. Protein sequencing analyses revealed that both complexes contained BUB3, a checkpoint protein involved in mitotic regulation. Immunoprecipitation and western blot analysis along with 35S-methionine labeling experiments, GST-pull down experiments, and gel filtration experiments confirmed the findings of a novel sumoylated 105kDa complex containing BUB3, SUMO-1 and C-JUN. Formation of the 105kDa complex is likely to be via covalent bonds. It is upregulated in cells approaching crisis, which is characterized by a mitotic arrest, and remains elevated in cells that have spontaneously recovered from such arrest, which are typically near tetraploid. Immortal ovarian carcinoma cell lines that are near tetraploid, including ML9A, SKOV3, and CAOV3, show detectable amounts of this complex, while cell lines that are near diploid, such as HEY and HOC-7, do not. Treatment with siRNA against BUB3 leads to a decrease in the 105kDa complex as well as native BUB3. Treatment of the cells with paclitaxel, a microtubule inhibitor, leads to intracellular accumulation of the complex, while treatment with aphidicolin, an S phase inhibitor, leads to its decrease, suggesting that formation of this complex is cell cycle dependent, occurring later than the S phase and preceding microtubule assembly.

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University of Southern California Dissertations and Theses

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Asset Metadata

Creator Lee, Jennifer Yeh (author)

Core Title The discovery of a novel 105kDa complex comprised of BUB3, C-JUN and SUMO-1 and its relationship to ovarian cancer development

School Keck School of Medicine

Degree Doctor of Philosophy

Degree Program Pathobiology

Publication Date 11/03/2009

Defense Date 10/09/2009

Publisher University of Southern California (original), University of Southern California. Libraries (digital)

Tag aneuploidy,BUB3,c-Jun,mitotic checkpoint,OAI-PMH Harvest,SUMO-1,tetraploidy

Language English

Contributor Electronically uploaded by the author (provenance)

Advisor Dubeau, Louis (committee chair), Ladner, Robert D. (committee member), Schonthal, Axel (committee member), Widelitz, Randall B. (committee member)

Creator Email jenny.yeh@gmail.com,jensyeh@hotmail.com

Permanent Link (DOI) https://doi.org/10.25549/usctheses-m2705

Unique identifier UC1300842

Identifier etd-Lee-3352 (filename),usctheses-m40 (legacy collection record id),usctheses-c127-271059 (legacy record id),usctheses-m2705 (legacy record id)

Legacy Identifier etd-Lee-3352.pdf

Dmrecord 271059

Document Type Dissertation

Rights Lee, Jennifer Yeh

Type texts

Source University of Southern California (contributing entity), University of Southern California Dissertations and Theses (collection)

Repository Name Libraries, University of Southern California

Repository Location Los Angeles, California

Repository Email cisadmin@lib.usc.edu