Chromodomain helicase DNA-binding (CHD) proteins are ATP-dependent chromatin remodeler enzymes that recognize methyl groups in lysines and participate in gene activation and repression, dependent on their cooperation with other chromatin reader complexes. The CHD family is composed of nine members (CDH1 to CHD9) grouped into three subfamilies based on the presence of additional functional domains. My group is interested in this family of enzymes because we found that CHD7 and CHD8 are mutated in 50% of CIMP1 colorectal carcinomas (CRCs). Although knowingly associated to developmental diseases, new studies have implicated these genes in other cancer types besides CRCs, including lung, gastric, and pancreatic tumors. Other CHD members are validated tumor suppressors, as examples CHD1 and CHD5, reinforcing the importance of the CHD family to tumor biology. In this study we will focus on the roles of CHD7 in epigenetic regulation with the goals of identifying directly regulated target genes and protein complexes that cooperate with this enzyme in normal and colon cancer. Colon cancer is third most deadly cancer type in the United States, and the premise of our research is that revealing facets of its epigenetic regulation will contribute positively to our understand of the disease and promote the development of therapies. We have collected preliminary data supporting that depletion of CHD7 results in increase in H3K27me3 levels, and deregulated the expression of known targets of the PRC1 and SNF2 complex. We will accomplish the goals of the project in two aims, using as models a collection of normal-derived, non-tumorigenic cell lines and colorectal carcinoma cell lines for which CHD7 down-regulation and up-regulation was manipulated by shRNAs and cDNA plasmids.
In aim 1 we will evaluate changes in gene expression that occur in response of altered expression of CHD7. We will also measure the effects of altered expression of CHD7 on DNA methylation and selected histone tails posttranslational modifications. Histone marks associated with enhancers and promoters are of key interest.
In aim 2, we will perform immunoprecipitation to validate and identify the interacting complexes that work along CHD7 in colon cells. Candidate partners are selected members of the PBAF and PRC1 complexes. CHD8 has been shown to act downstream to BRAF mutations to mediate DNA hypermethylation of CIMP genes, and we will also evaluate whether CHD7 partners with CHD8 and DNMT3A/B in wild type and mutated BRAF CRCs. Mass spectrometry will be used to account for unexpected interactions. The results of this project are expected to bring novel insights into how CHD7 in particular, and epigenetic alterations in general contribute to colorectal carcinomas, and identify novel pathways involved in CRC tumorigenesis that may represent opportunities for therapeutic intervention. In addition, they will also provide basic information on the role of CHD7 in gene regulation and its interacting complexes in colon cells.
Genes that regulate the epigenome represent new promising therapeutic targets, and the development of drugs that target this class of enzymes can be improved by a better understanding of the functions of these enzymes and the roles they play in cancer, including the identification of downstream targets with key roles in the disease. As a group, chromatin-remodeling genes are found frequently mutated in multiple tumor types; our own research identified CHD7 as frequently mutated in a specific subgroup of colorectal carcinomas. In this project we will investigate the role of this chromatin-remodeling enzyme in the regulation of the epigenome and selected signaling pathways in colon, and its relationship to abnormal epigenetic states in colorectal carcinomas.
