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BRCA2 and PALB2 in Cellular Stress Response

by
Allen Alcivar
B.S., Kean University - 2007

Thesis Advisors: Bing Xia, Ph.D.
Graduate Program in Cellular & Molecular Pharmacology

Cancer Institute of New Jersey
Auditorium A
New Brunswick

Thursday, August 7, 2014
1:00 p.m.


Abstract

Mutations in the BRCA2 (Breast Cancer 2, early onset) gene are implicated in a variety of familial cancers. About 50% of BRCA2 is associated with PALB2 (partner and localizer of BRCA2), which anchors BRCA2 onto chromatin and directs its recruitment to DNA damage sites. Loss of BRCA2 function results in severe defects in DNA double-strand break repair, DNA damage-induced checkpoint response and the stability of stalled DNA replication forks, all of which are important for genomic stability. How BRCA2 mediates checkpoint response and stalled replication fork stability is poorly understood.
We report an interaction between BRCA2 and an essential replication factor that promotes initiation of DNA replication, MCM10. Our findings demonstrate that MCM10 and BRCA2 interact through specific motifs. Cells depleted of BRCA2 or MCM10 showed similar instability of stalled replication forks. Moreover, DNA damage-induced CHK1 activation is markedly reduced in both BRCA2- and MCM10-depleted cells even though RPA is hyperphosphorylated. Our studies suggest BRCA2-MCM10 interaction may be important for checkpoint control as well as stalled replication fork stability.
In addition, we propose BRCA2 directs the localization and expression of an important DNA replication licensing factor, CDC6. This regulation of CDC6 is exclusive from other hereditary breast cancer-associated proteins, namely BRCA1 and PALB2. In particular, CDC6 chromatin association and localization is misregulated in cells knocked down for BRCA2 expression. Patient-derived cells with nonfunctional BRCA2 also show aberrant CDC6 localization and expression. These data suggest BRCA2 function in checkpoint control may be mediated by CDC6.
Finally, we present another mechanism that promotes genomic stability through maintenance of proper oxidative stress response. Cells that cannot efficiently mitigate reactive oxygen species (ROS) are susceptible to mutations and carcinogenesis. Kelch-like ECH-associated Protein 1 (KEAP1) is the main negative regulator of the NFE2-related Factor 2 (NRF2)-driven oxidative stress response. Earlier studies from our lab demonstrate PALB2 interacts with KEAP1 and can regulate NRF2 function. Our present results identify Dipeptidyl Peptidase III (DPP3) as a significant binding partner of KEAP1 upon oxidative stress induction. Overexpression of DPP3 promotes NRF2 nuclear accumulation and transcriptional activity in addition to conferring resistance to oxidative stress. These functions are dependent on DPP3-KEAP1 interaction and indicate DPP3 is an important mediator of normal oxidative stress response.


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