A High Resolution Genome-Wide Scan of HNF4α Recognition Sites Infers a Regulatory Gene Network in Colon Cancer

The hepatic nuclear factor HNF4 alpha is a versatile transcription factor and controls expression of many genes in development, metabolism and disease. To delineate its regulatory gene network in colon cancer and to define novel gene targets a comprehensive genome-wide scan was carried out at a resolution of 35 bp with chromatin IP DNA obtained from the human colon carcinoma cell line Caco-2 that is a particularly rich source of HNF4 alpha. More than 90% of HNF4 alpha binding sites were mapped as promoter distal sequences while enhancer elements could be defined to foster chromatin loops for interaction with other promoter-bound transcription factors. Sequence motif analysis by various genetic algorithms evidenced a unique enhanceosome that consisted of the nuclear proteins ER alpha, AP1, GATA and HNF1 alpha as cooperating transcription factors. Overall >17,500 DNA binding sites were identified with a gene/binding site ratio that differedw >6-fold between chromosomes and clustered in distinct chromosomal regions amongst >6600 genes targeted by HNF4 alpha. Evidence is presented for nuclear receptor cross-talk of HNF4 alpha and estrogen receptor a that is recapitulated at the sequence level. Remarkably, the Y-chromosome is devoid of HNF4 alpha binding sites. The functional importance of enrichment sites was confirmed in genome-wide gene expression studies at varying HNF4 alpha protein levels. Taken collectively, a genome-wide scan of HNF4 alpha binding sites is reported to better understand basic mechanisms of transcriptional control of HNF4 alpha targeted genes. Novel promoter distal binding sites are identified which form an enhanceosome thereby facilitating RNA processing events.