Growth Hormone-Activated STAT5 May Indirectly Stimulate IGF-I Gene Transcription through HNF-3γ

Abstract

IGF-I is abundantly expressed in the liver under the stimulation of GH. We showed previously that expression of hepatocyte nuclear factor (HNF)-3γ, a liver-enriched transcription factor, was strongly stimulated by GH in bovine liver. In this study, we determined whether GH-increased HNF-3γ might contribute to GH stimulation of IGF-I gene expression in bovine liver and the underlying mechanism. A sequence analysis of the bovine IGF-I promoter revealed three putative HNF-3 binding sites, which all appear to be conserved in mammals. Chromatin immunoprecipitation assays showed that GH injection increased binding of HNF-3γ to the IGF-I promoter in bovine liver. Gel-shift assays indicated that one of the three putative HNF-3 binding sites, HNF-3 binding site 1, bound to the HNF-3γ protein from bovine liver with high affinity. Cotransfection analyses demonstrated that this HNF-3 binding site was essential for the transcriptional response of the IGF-I promoter to HNF-3γ in CHO cells and to GH in primary mouse hepatocytes. Using similar approaches, we found that GH increased binding of the signal transducer and activator of transcription 5 (STAT5) to the HNF-3γ promoter in bovine liver, that this binding occurred at a conserved STAT5 binding site, and that this STAT5 binding site was necessary for the HNF-3γ promoter to respond to GH. Taken together, these results suggest that in addition to direct action, GH-activated STAT5 may also indirectly stimulate IGF-I gene transcription in the liver by directly enhancing the expression of the HNF-3γ gene.