RFX1 and NF-1 associate with P sequences of the human growth hormone locus in pituitary chromatin

The human GH family consists of five genes, including the placental chorionic somatomammotropins ( CS), within a single locus on chromosome 17. Based on nuclease sensitivity, the entire GH/CS locus is accessible in pituitary chromatin, yet only GH-N is expressed. Previously, we reported a P sequence element (263P) capable of repressing placental CS promoter activity in transfected pituitary ( GC) cells. Regions of protein binding within 263P include P sequence elements A and B (PSE-A and PSE-B), and we reported nuclear factor-1 (NF-1) recognition of PSE-B. We now provide evidence for multiple interactions on PSE-A, including binding of the regulatory factor X (RFX) family. Disruption of the RFX site within 263P blunts repressor activity in transfected GC cells; however, repression is only abolished when both PSE-A/RFX and PSE-B/NF-1 sites are mutated. The capacity of RFX and NF-1 to participate in a novel common complex is further suggested by coimmunoprecipitation of RFX1 and epitope-tagged NF-1 family members. Finally, we confirm the association of NF-1 and RFX1 with P sequences in human pituitary tissue by chromatin immunoprecipitation. Taken together, our data suggest that an inverse relationship exists between 263P and CS promoter histone hyperacetylation and the association of these factors in vivo.