Identification and functional analysis of a locus control region for the human growth hormone gene

The human growth hormone/chorionic somatomammotropin (hGH/hCS) gene cluster contains five highly homologous genes displaying mutually exclusive expression in two tissues. hGH-N is expressed solely in the pituitary; transcription of hCS-L, hCS-A, hCS-B, and hGH-V is restricted to the placenta. hGH-N, hCS-A, and hCS-B are expressed at extremely high levels in vivo; however hGH-N and hCS-A, when accompanied by all previously defined regulatory sequences, are not expressed well after introduction into the germline of transgenic mice. Mice often fail to express these genes; only low levels of transgene transcription are detected in expressing mice, suggesting that these genes are subject to positional effects, governed by where they integrate into the host genome. In order to identify additional regulatory elements, that would consistently direct high-level expression of the hGH/hCS genes, DNase I hypersensitive sites were mapped in pituitary and placental chromatin. 40kb of 5$\sp\prime$ flanking sequence, that includes all identified 5$\sp\prime$ DNase I hypersensitive sites, consistently directs high-level, position-independent, copy number-dependent expression of hGH-N in the transgenic mouse pituitary. Two different hypersensitive site containing regions mediate position-independent hGH-N expression in the pituitary, one of which strongly enhances hGH-N expression in the transgenic mouse pituitary but is relatively inactive in transfected rat pituitary cells. Neither hypersensitive site region alone functions as effectively as the intact 40kb 5$\sp\prime$ flanking domain. One hypersensitive site region, when linked directly to the hGH-N gene, consistently directs high-level expression of the transgene in the kidney, suggesting that synergistic interactions between regulatory sequences are required to restrict hGH-N transcription to the pituitary. These results establish the presence of a locus control region for the hGH-N gene cluster, and indicate that full activity requires the cooperative interaction of multiple regulatory elements