A243 FUNCTIONAL ROLES OF NCOR1 AND CHD8 PROTEIN INTERACTION IN HUMAN COLORECTAL CANCER CELLS. (1st March 2018) - Loiselle, A
Abstract: Background: The nuclear receptor co-repressor 1 (NCOR1) is a protein with transcriptional repression activity interacting with nuclear receptors. NCOR1 is involved in targeting gene transcription related to intestinal inflammatory response and colorectal cancer. We have previously identified the chromatin helicase DNA binding 8 (CHD8) protein as a potential interactor of NCOR1. CHD8 is a DNA helicase that functions as a transcription repressor by remodeling chromatin structure. Aims: To investigate the functional and biological roles of CHD8 interaction with NCOR1 in colorectal cancer. Methods: HT-29 and Caco-2/15 cell lines were depleted in NCOR1 or CHD8 by RNAi. Cell proliferation of the generated cell lines was measured by cell counts. Overall changes in transcriptome was determined by high-throughput RNA sequencing. Chromatin immunoprecipitation (ChIP) was performed with the use of a specific CHD8 antibody. Multiple HA and V5 epitope-tagged domain fragments of NCOR1 and CHD8 were generated for interaction assays. Results: Depletion of NCOR1 in HT-29 and Caco-2/15 cells led to a strong reduction of cell proliferation, while depletion of CHD8 led to a less robust effect. High-throughput RNA sequencing identified more than 60 common genes for which the expression was modulated in both NCOR1 and CHD8 depleted HT-29 and Caco-2/15 cell populations. Among these modulated genes, the Brain Derived Neutrophic Factor (BDNF), known to be involved in cancer cell motility, was identified as a common gene target for both NCOR1 and CHD8. ChIP with the use of a CHD8 specific antibody was optimized in colorectal cancer cell lines to monitor the level of CHD8 chromatin occupancy among the identified genes. Finally, various tagged constructs representative of specific domains of NCOR1 and CHD8 have been generated and validated by Western blot. Interactions strategies are currently undergoing to functionally validate NCOR1 and CHD8 biological link. Conclusions: NCOR1 and CHD8 interact together, affect common genes and negatively influence colorectal cancer cell proliferation. Identification of the specific nature of this interaction will highlight novel strategies to disrupt NCOR1-CHD8 interaction in order to regulate colorectal cancer cell proliferation. Funding Agencies: CIHR