Talk abstracts

Talk on Monday 03:30-03:45pm submitted by Shweta Kotian

Histone Deacetylases 9 and 10 are Required for Homologous Recombination

Shweta Kotian (Department of Biomedical Informatics, The Ohio State University Comprehensive Cancer Center), Sandhya Liyanarachchi (Biomedical Informatics Shared Resource, The Ohio State University Comprehensive Cancer Center), Arthur Zelent (Section of Haemato-Oncology, Institute for Cancer Research, Sutton, Surrey), Jeffrey D. Parvin (Department of Biomedical Informatics, The Ohio State University Comprehensive Cancer)

Abstract:
Histone Deacetylases (HDACs) along with histone acetylases (HATs) are known to be involved in the regulation of a wide spectrum of factors involved in transcription, the cell cycle, development, differentiation etc. The former class leads to chromatin compaction, which is transcriptionally repressive, while the latter leads to open chromatin that stimulates transcription. This led to our initial hypothesis that inhibition of HDACs would allow for better access to damaged chromatin by repair factors, thereby, stimulating DNA repair processes. In addition, HDAC inhibitors have attracted significant research interest as cancer therapeutics, but not much is known about their exact mechanism of action. Therefore, we set out to test the role of HDACs in the homologous recombination process. A tissue-culture based homology directed repair (HDR) assay was used in which repair of a double-stranded break, by homologous recombination, results in gene conversion of an inactive GFP allele to an active GFP gene. Homologous recombination events were then readily scored by measuring GFP positive cells by FACS. The HDAC inhibitors used were trichostatin A (TSA), apicidin, valproic acid (VPA) and sodium butyrate (NaB). Contrary to our hypothesis, treatment of cells with the inhibitors significantly reduced homologous recombination upto 11-fold. Using RNA interference to deplete each HDAC, we found that HDAC9 and HDAC10 depletion specifically inhibited homologous recombination. By assaying for sensitivity of cells to the inter-strand crosslinker mitomycin C, we found, that either treatment of cells with HDAC inhibitors or depletion of HDAC9 or HDAC10, resulted in increased sensitivity to mitomycin C, consistent with the HDR assay results. In conclusion, HDAC9 and HDAC10 are required for the homologous recombination process. We believe that these enzymes catalyze an important deacetylation step at the site of DNA damage. Current efforts are directed toward finding the crucial substrate.

Keywords: DNA repair, deacetylases, cancer therapy