Type of Document	Dissertation
Author	Liu, Hanjian
Author's Email Address	hanjian.liu@vanderbilt.edu
URN	etd-09152007-110210
Title	HUMAN DNA HELICASE B FUNCTIONS IN DNA DAMAGE RESPONSE AND HOMOLOGOUS RECOMBINATION
Degree	PhD
Department	Biological Sciences
Advisory Committee	Advisor Name Title James Patton Committee Chair David Cortez Committee Member Ellen Fanning Committee Member Eugene Oltz Committee Member Jennifer Pietenpol Committee Member
Keywords	ATR homologous recombination helicase DNA damage response chromatin association DNA repair
Date of Defense	2007-05-23
Availability	unrestricted
Abstract DNA damaging agents have been shown to stimulate the localization of human DNA helicase B (HDHB) in nuclear foci, suggesting that HDHB might participate in DNA damage response. In the first part of this dissertation, we found that the chromatin-associated fraction of HDHB increases in cells exposed to a variety of DNA damaging agents. HDHB chromatin accumulation is most prominent in S phase cells exposed to agents that cause replication fork stalling or collapse. Inhibition of checkpoint kinases does not prevent damage-induced accumulation of HDHB on chromatin, suggesting that HDHB associates directly with DNA lesions or with other proteins recruited to lesions. Silencing of HDHB does not affect UV-induced RPA focus formation, but diminishes induction of TopBP1 foci. DNA damage induced CHK1 phosphorylation is impaired in HDHB-depleted cells. These results identify HDHB as a novel factor that associates with damaged chromatin and promotes intra-S phase damage responses. In the second part of this dissertation, we further investigated the possible function of HDHB in homologous recombination. HDHB-depleted cells showed more aphidicolin-induced chromosome breaks than control-depleted cells. HDHB-depleted cells have fewer sister chromatid exchange than control-depleted cells. An in vivo recombination assay showed that HDHB silencing results in impaired homologous recombination. In vitro, recombinant HDHB stimulates Rad51-mediated 5’-3’ heteroduplex extension. Our studies suggest a function of HDHB in stimulating ssDNA/duplex structure during homologous recombination.
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