Expression of the Hepatic Specific V1 Messenger Ribonucleic Acid of the Human Growth Hormone Receptor Gene Is Regulated by Hepatic Nuclear Factor (HNF)-4α2 and HNF-4α8

Goodyer, Cynthia G.; Rhani, Zakaria; Zheng, Hong

doi:10.1210/me.2007-0387

Cited by 11 publications

(3 citation statements)

References 62 publications

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“…The largest V2(Ϫ2623/ϩ335) construct was created by three steps: 1) a 3100-bp fragment was PCR amplified using forward primer V2F(Ϫ2623) and reverse primer V2R(Ϫ512) (supplemental Table 1, published as supplemental data on The Endocrine Society's Journals Online web site at http://mend.endojournals.org) with human genomic DNA as a template; 2) this 3100-bp fragment was cloned into pCR2.1-TOPO vector (Invitrogen); and 3) the MluI-BsrBI fragment containing the V2(Ϫ2623/ϩ335) piece was excised and ligated into pGL3-basic using MluI and SmaI sites. The V2(Ϫ721/ϩ362) construct was engineered by HindIII digestion of a plasmid containing the hGHR V2 exon and approximately 1 kb of 5Ј-upstream region (21) and ligated into the HindIII site of pGL3-basic. The V2(Ϫ211/ϩ362) and V2(ϩ11/ϩ362) reporter constructs were prepared by PCR amplification and subcloned into pGL3-basic.…”

Section: Generation Of V2 Promoter Deletion and Mutation Reporter Conmentioning

confidence: 99%

“…The hGHR is encoded by a single gene on chromosome 5p13.1-p12 that contains several 5Ј-untranslated exons under the control of different promoters (15)(16)(17)(18)(19)(20)(21). The multiple mRNA variants (V) transcribed from the different 5Ј-untranslated region (5Ј-UTR) noncoding exons splice into a common acceptor site in the first coding exon, 11 bp upstream of the ATG translation start codon; thus, all transcripts encode the same hGHR protein.…”

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confidence: 99%

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Structure and Activity of the Human Growth Hormone Receptor (hGHR) Gene V2 Promoter

Wei¹,

Puzhko²,

Wabitsch³

et al. 2009

Molecular Endocrinology

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Human GH (hGH) has important effects on growth as well as carbohydrate, fat, and protein metabolism. These actions require the presence of normal levels of a functional hGH receptor (hGHR) on the surface of target cells. hGHR gene expression is characterized by the use of several 5'-noncoding exons and alternative splicing, resulting in the generation of multiple mRNA isoforms. The hGHR V2 transcript is predominant in most tissues, including human fat. However, factors regulating its ubiquitous expression have remained unidentified. The present study was aimed at characterizing the mechanisms regulating hGHR V2 transcription. Two major V2 transcriptional start sites were identified by primer extension assays. The V2 proximal promoter is TATA-less, with several characteristics of a housekeeping gene promoter. Transient transfection analyses of 2.6 kb of the 5'-flanking region of V2 confirmed its promoter activity in multiple primate cell lines. Similar promoter activity patterns were observed in human SGBS preadipocytes and mature adipocytes but with much higher V2 promoter activity in mature adipocytes, suggesting that changes in the availability of specific factors during adipocyte differentiation play a role in V2 promoter regulation. Serial deletion and mutation analyses revealed that transcription of hGHR V2 in different cell types, including adipocytes, is determined by a core promoter and distinct inhibitory and activation domains in the 5'-promoter region as well as within the V2 exon. Our data suggest that V2 transcription is the result of a complex interplay involving multiple factors, to ensure appropriate expression of hGHR in different hGH target cells.

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Section: Generation Of V2 Promoter Deletion and Mutation Reporter Conmentioning

confidence: 99%

mentioning

confidence: 99%

Structure and Activity of the Human Growth Hormone Receptor (hGHR) Gene V2 Promoter

Wei¹,

Puzhko²,

Wabitsch³

et al. 2009

Molecular Endocrinology

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“…There have been extensive studies of how GHR gene expression is regulated at its multiple 5ЈUTR promoters by our lab (15)(16)(17)(18) as well as others (19). However, potential regulation at the 3ЈUTR has not been examined.…”

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Regulation of Human Growth Hormone Receptor Expression by MicroRNAs

Elzein

Goodyer

2014

Molecular Endocrinology

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Human GH binds to its receptor (GHR) on target cells and activates multiple intracellular pathways, leading to changes in gene expression, differentiation, and metabolism. GHR deficiency is associated with growth and metabolic disorders whereas increased GHR expression has been reported in certain cancers, suggesting that the GHR gene requires tight controls. Several regulatory mechanisms have been found within its 5'-untranslated region (UTR) promoter and coding regions. However, the 3'-UTR has not been previously examined. MicroRNAs (miRNAs) are small (19-22 nucleotides) noncoding RNAs that downregulate gene expression mainly through targeting the 3'-UTR of mRNAs and enhancing their degradation or inhibiting translation. In the present study, we investigated whether miRNAs regulate GHR expression. To define putative miRNA binding sites in the GHR 3'-UTR, we used multiple in silico prediction tools, analyzed conservation across species and the presence of parallel sites in GH/IGF axis-related genes, and searched for reports linking miRNAs to GHR-related physiological or pathophysiological activities. To test prioritized sites, we cotransfected a wild-type GHR 3'-UTR luciferase reporter vector as well as miRNA binding site mutants into HEK293 cells with miRNA mimics. Furthermore, we tested whether the miRNAs altered endogenous GHR mRNA and protein levels in HEK293 cells and in 2 cancer cell lines (MCF7 and LNCaP). Our experiments have identified miRNA (miR)-129-5p, miR-142-3p, miR-202, and miR-16 as potent inhibitors of human GHR expression in normal (HEK293) and cancer (MCF7 and LNCaP) cells. This study paves the way for the development of miRNA inhibitors as therapeutic agents in GH/GHR-related pathophysiologies, including cancer.

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