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MITOTIC SLIPPAGE IN NONCANCER CELLS INDUCED BY A MICROTUBULE DISRUPTOR, DISORAZOLE C1, AND THE NOVEL ROLE FOR GPIBA IN CYTOKINESIS AND TETRAPLOIDIZATION IN CANCER CELLS

Xu, Fengfeng (2010) MITOTIC SLIPPAGE IN NONCANCER CELLS INDUCED BY A MICROTUBULE DISRUPTOR, DISORAZOLE C1, AND THE NOVEL ROLE FOR GPIBA IN CYTOKINESIS AND TETRAPLOIDIZATION IN CANCER CELLS. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Abstract

Disorazole C1 (DZ) is a synthesized polyene macrodiolide, originally identified as a minor metabolite from the myxobacterium fermentation broth. I examined the cellular responses to DZ in noncancer and cancer cells, and compared the results to vinblastine and taxol. In noncancer cells, DZ induced a prolonged mitotic arrest, followed by mitotic slippage, which was associated with cyclin B degradation, but did not require p53. Four apoptosis detection assays, including examination of poly(ADP-ribose) polymerase cleavage, cytochrome C release, mitochondrial depolarization, and annexin V staining, were conducted and demonstrated little induction of apoptosis in noncancer cells treated with DZ. However, I observed an activated apoptotic pathway in cancer cells, suggesting that normal and malignant cells respond differently to DZ, and indicating a potential therapeutic application for this compound.GpIba is a transmembrane subunit of the von Willebrand factor receptor on platelet surfaces, functioning in platelet adhesion and activation. Recent research has revealed that GpIba also plays roles in transformation and genomic instability as an oncoprotein. In my studies, GpIba colocalized with F-actin and filamin A, an actin-binding protein, at the cleavage furrow in anaphase cells, suggesting a novel role for GpIba in cytokinesis. However, when GpIba was overexpressed in noncancer cells with p53 knockdown, GpIba, F-actin, and filamin A became mislocalized. RhoA, a cytokinesis regulator, was localized asymmetrically, although Aurora B, an important cytokinesis kinase, retained its correct localization. Additionally, anaphase bridges were observed in these GpIba-overexpressing cells, along with elevated percentages of cytokinesis failure and binucleation, indicating that GpIba overexpression led to cytokinesis defects/failure and tetraploidization. Conversely, in cancer cells that endogenously overexpress GpIba, I observed decreases in the percentages of binucleation and mitotic defects upon GpIba knockdown. Furthermore, down-regulation of Aurora B was demonstrated to mediate the mechanism through which GpIba overexpression led to cytokinesis failure, and the filamin A-binding domain and the signal peptide of GpIba were shown to be indispensible for this mechanism. These results add a new component of the existing cancer cell genetic instability pathways, by suggesting that overexpression of GpIba in cancer cells disrupts normal cytokinesis and causes tetraploidization through down-regulation of Aurora B.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Xu, Fengfengffxuustc@gmail.com
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairSaunders, William Swsaund@pitt.eduWSAUND
Committee MemberDay, Billy Wbday@pitt.eduBDAY
Committee MemberPipas, James Mpipas@pitt.eduPIPAS
Committee MemberKiselyov, Kirillkiselyov@pitt.eduKISELYOV
Committee MemberJacobson, Lewis Aljac@pitt.eduLJAC
Date: 24 June 2010
Date Type: Completion
Defense Date: 25 March 2010
Approval Date: 24 June 2010
Submission Date: 15 April 2010
Access Restriction: 5 year -- Restrict access to University of Pittsburgh for a period of 5 years.
Institution: University of Pittsburgh
Schools and Programs: Dietrich School of Arts and Sciences > Biological Sciences
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: apoptosis; cancer; cell division; cytokinesis; disorazole; GpIba; mitosis; mitotic slippage; tumor
Other ID: http://etd.library.pitt.edu/ETD/available/etd-04152010-160048/, etd-04152010-160048
Date Deposited: 10 Nov 2011 19:37
Last Modified: 15 Nov 2016 13:40
URI: http://d-scholarship.pitt.edu/id/eprint/7205

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