Double strand break (DSB) processing has important implications for genomic instability, radio- sensitivity and cancer. Recent studies have shed light on the significance of chromatin remodeling in relation to DSBs as well as ubiquitin-dependent transcriptional regulation at these sites. The ND10 nuclear body is a multi-protein nuclear structure that has been shown to recognize DSBs and engage in transcriptional silencing of viral genomes. Composed of SUMOylated protein components PML, Sp100 and others, ND10 bodies are also the target of degradation by ICP0, a viral protein produced during HSV-1 infection to counteract ND10 silencing mechanisms and promote viral genome transcription and replication. Preliminary data show that SUMOylation plays a role in transcriptional silencing near DSB foci as well, as expression of ICP0 strongly reverses silencing at these foci. ND10 bodies are also recruited to hypomethylated and decondensed pericentromeric heterochromatin in a disorder known as immunodeficiency, centromere instability and facial anomalies (ICF) syndrome. Transcription from these regions as well as other repetitive elements such as retrotransposons have been observed to occur in normal cells that are stressed or exposed to DNA damaging agents. In addition, overexpression of satellite transcripts has been observed in human pancreatic ductal adenocarcinomas as well as in other epithelial cancers. The purpose of this project is to determine the manner in which SUMOylation mediates the transcriptional silencing associated with the seemingly disparate realms of DNA damage, viral genomes and constitutive heterochromatin. Immunofluorescence studies involving satellite specific probes will be conducted to determine the persistence of IR-induced foci (IRIF), their localization within the nucleus, and the significance of satellite transcription on both of these aspects of IRIF. The significance of SUMOylation on transcriptional repression at sites of DSBs will be examined using a novel system that induces DSBs at endogenous sites of the genome. Transcription of genes in cis to these regions will be measured using quantitative PCR, and the presence of SUMO moieties at promoter regions of genes located near DSBs will be assessed by chromatin immunoprecipitation. Similar studies will be conducted on viral genomes. Determining the function of SUMOylation, the response of cells to DSBs and heterochromatin, and a cell's innate anti-viral response could be informative in understanding their role in genomic instability and in the development of radio resistance that is observed in certain cancers.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Individual Predoctoral NRSA for M.D./Ph.D. Fellowships (ADAMHA) (F30)
Project #
5F30CA177190-02
Application #
8643479
Study Section
Special Emphasis Panel (ZRG1-F08-Q (20))
Program Officer
Damico, Mark W
Project Start
2013-04-01
Project End
2015-05-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
2
Fiscal Year
2014
Total Cost
$29,227
Indirect Cost
Name
University of Pennsylvania
Department
None
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104