Date of Award

Spring 2009

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biological Sciences

Program/Concentration

Biomedical Sciences

Committee Director

O. John Semmes

Committee Member

Richard Drake

Committee Member

Julie Kerry

Committee Member

Neel Krishna

Abstract

Human T-cell leukemia virus type 1 (HTLV-1) is a transforming retrovirus that can give rise to adult T-cell leukemia (ATL) and HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP). Tax is a virally encoded oncoprotein that is involved in HTLV-1 mediated cellular transformation. It has been hypothesized that Tax induces genomic instability via repression of the cellular DNA damage repair response. Our laboratory has previously shown that the interaction between Tax and various proteins involved in the DNA-damage response pathway impairs the ability of these proteins to mount an efficient repair response. As part of these observations, we proposed that Tax induces a constitutive phosphorylation state in the DNA-dependent Protein Kinase (DNA-PK) which effectively saturates the naive response to new DNA damage events.

In this study, using Liquid Chromatography-Mass Spectrometry, we determined that the mediator of DNA damage checkpoint protein 1 (MDC1) is also part of the Tax-bound complex. MDC1 is a DNA damage repair (DDR) protein that is found in nuclear foci called ionizing radiation-induced foci (IRIF) that form in response to DNA damage. In this study we show that MDC1, which is a part DNA-PK-MDC1 signaling pathway, is active in Tax expressing cells. In addition, we demonstrate that Tax recruits MDC1 to "Tax speckled structures" (TSS). The same observation was made when the localization of other DDR proteins such as DNA-PK were examined. These results suggest an ability of Tax to generate a damage-independent DDR response. Here, we demonstrate that MDC1 is recruited away from the TSS to the IRIF following damage, whereas DNA-PK phosphorylated at threonine 2609, an activated form of DNA-PK, and BRCA1 were shared between the two entities. These results suggest that there is a limited amount of MDC1 in the cell that is competed for between TSS and IRIF. One potential impact of generating a damage independent DDR response is competitive sequestering of components of the DDR away from the actual sites of damage. This competitive recruitment of cellular DDR may explain the observation that Tax expression results in genomic instability.

DOI

10.25777/n7v1-2a08

ISBN

9781109594195

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