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Growth hormone-activated STAT5 may indirectly stimulate IGF-I gene transcription through HNF-3{gamma}.

ABSTRACT: IGF-I is abundantly expressed in the liver under the stimulation of GH. We showed previously that expression of hepatocyte nuclear factor (HNF)-3gamma, a liver-enriched transcription factor, was strongly stimulated by GH in bovine liver. In this study, we determined whether GH-increased HNF-3gamma 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-3gamma 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-3gamma 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-3gamma 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-3gamma 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-3gamma 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-3gamma gene.

SUBMITTER: Eleswarapu S

PROVIDER: S-EPMC2796150 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature

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Growth hormone-activated STAT5 may indirectly stimulate IGF-I gene transcription through HNF-3{gamma}.

Eleswarapu Satyanarayana S Ge Xiaomei X Wang Ying Y Yu Jie J Jiang Honglin H

Molecular endocrinology (Baltimore, Md.) 20091009 12

IGF-I is abundantly expressed in the liver under the stimulation of GH. We showed previously that expression of hepatocyte nuclear factor (HNF)-3gamma, a liver-enriched transcription factor, was strongly stimulated by GH in bovine liver. In this study, we determined whether GH-increased HNF-3gamma 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, w ...[more]

PMID: 19819986

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Project description:Many of the physiological actions of GH are mediated by IGF-I, a secreted 70-residue peptide whose gene expression is induced by GH in the liver and other tissues via mechanisms that remain incompletely characterized but depend on the transcription factor Stat5b. Here we investigate the chromatin landscape of the IGF-I gene in the liver of pituitary-deficient young adult male rats and assess the impact of a single systemic GH injection. Despite minimal ongoing transcription in the absence of GH, both IGF-I promoters appear to reside in open chromatin environments, at least as inferred from relatively high levels of acetylation of core histones H3 and H4 when compared with adjacent intergenic DNA and from enhanced trimethylation of histone H3 at lysine 4. This landscape of open chromatin may reflect maturation of the liver. Surprisingly, in the absence of hormone, IGF-I promoter 1 appears poised to be activated, as evidenced by the presence of the transcriptional coactivator p300 and recruitment of RNA polymerase (Pol) II into a preinitiation complex. By contrast, chromatin surrounding IGF-I promoter 2 is devoid of both p300 and RNA Pol II. Systemic GH treatment causes an approximately 15-fold increase in transcription from each IGF-I promoter within 60 min of hormone administration, leading to a sustained accumulation of IGF-I mRNA. The coordinated induction of both IGF-I promoters by GH is accompanied by hyperacetylation of histones H3 and H4 in promoter-associated chromatin, a decline in monomethylation at lysine 4 of histone H3, and recruitment of RNA Pol II to IGF-I promoter 2. We conclude that GH actions induce rapid and dramatic changes in hepatic chromatin at the IGF-I locus and activate IGF-I gene transcription in the liver by distinct promoter-specific mechanisms: at promoter 1, GH causes RNA Pol II to be released from a previously recruited paused preinitiation complex, whereas at promoter 2, hormone treatment facilitates recruitment and then activation of RNA Pol II to initiate transcription.

Project description:Growth hormone (GH) facilitates therapy resistance in the cancers of breast, colon, endometrium, and melanoma. The GH-stimulated pathways responsible for this resistance were identified as suppression of apoptosis, induction of epithelial-to-mesenchymal transition (EMT), and upregulated drug efflux by increased expression of ATP-binding cassette containing multidrug efflux pumps (ABC-transporters). In extremely drug-resistant melanoma, ABC-transporters have also been reported to mediate drug sequestration in intracellular melanosomes, thereby reducing drug efficacy. Melanocyte-inducing transcription factor (MITF) is the master regulator of melanocyte and melanoma cell fate as well as the melanosomal machinery. MITF targets such as the oncogene MET, as well as MITF-mediated processes such as resistance to radiation therapy, are both known to be upregulated by GH. Therefore, we chose to query the direct effects of GH on MITF expression and activity towards conferring chemoresistance in melanoma. Here, we demonstrate that GH significantly upregulates MITF as well as the MITF target genes following treatment with multiple anticancer drug treatments such as chemotherapy, BRAF-inhibitors, as well as tyrosine-kinase inhibitors. GH action also upregulated MITF-regulated processes such as melanogenesis and tyrosinase activity. Significant elevation in MITF and MITF target gene expression was also observed in mouse B16F10 melanoma cells and xenografts in bovine GH transgenic (bGH) mice compared to wild-type littermates. Through pathway inhibitor analysis we identified that both the JAK2-STAT5 and SRC activities were critical for the observed effects. Additionally, a retrospective analysis of gene expression data from GTEx, NCI60, CCLE, and TCGA databases corroborated our observed correlation of MITF function and GH action. Therefore, we present in vitro, in vivo, and in silico evidence which strongly implicates the GH-GHR axis in inducing chemoresistance in human melanoma by driving MITF-regulated and ABC-transporter-mediated drug clearance pathways.

Project description:The transcriptional repressor Bcl6 is a male-specific rat liver gene product and one of 24 early GH-response genes encoding DNA-binding proteins. Presently, the sex specificity of Bcl6 was shown to emerge at puberty, when hepatic Bcl6 mRNA was induced in males and repressed in females by the female plasma GH profile. Hepatic Bcl6 mRNA was increased to near-normal male levels in hypophysectomized females and was extinguished in intact males given a continuous GH infusion (female-like GH pattern). Bcl6 was also repressed in adult male somatostatin-deficient mice, where plasma GH profiles are female like. Hepatic Bcl6 RNA was rapidly down-regulated by GH pulse treatment, both in hypophysectomized male rats and in primary rat hepatocytes. Bcl6 was substantially induced in female mice deficient in hepatic signal transducer and activator of transcription (STAT)5a/STAT5b, suggesting that these STAT transcriptional mediators of GH signaling repress Bcl6. Indeed, STAT5 was bound to Bcl6 STAT5-binding region-B, previously associated with Bcl6 repression, in both male and female liver chromatin. STAT5 also bound to Bcl6 region-A in male chromatin but only during a plasma GH pulse. Analysis of primary transcripts (heterogeneous nuclear RNA) across the Bcl6 gene revealed a novel mechanism of GH-dependent sex specificity, with two apparent blocks in Bcl6 transcription elongation seen in female liver and in continuous GH-treated male liver, one early in intron 4 and one in exon 5, which together reduced transcription beyond exon 5 more than 300-fold. Finally, Bcl6 was bound to a subset of STAT5-binding sites in male liver chromatin, including a Socs2 STAT5-binding site where Bcl6 binding increased substantially between plasma GH pulses, i.e. when STAT5 binding was low. Bcl6 and STAT5 binding are thus inversely coordinated by the endogenous pulses of pituitary GH release, suggesting this male-specific transcriptional repressor modulates hepatic GH signaling to select STAT5 target genes.

Dataset Information

Growth hormone-activated STAT5 may indirectly stimulate IGF-I gene transcription through HNF-3{gamma}.

Publications

Growth hormone-activated STAT5 may indirectly stimulate IGF-I gene transcription through HNF-3{gamma}.

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