Browsing by Author "Ge, Xiaomei"

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    Growth Hormone Stimulates Transcription of the Fibroblast Growth Factor 21 Gene in the Liver through the Signal Transducer and Activator of Transcription 5
    Yu, Jie; Zhao, Lidan; Wang, Aihua; Eleswarapu, Satyanarayana; Ge, Xiaomei; Chen, Daiwen; Jiang, Honglin (Endocrine Society, 2012-02)
    Fibroblast growth factor 21 (FGF21) is a recently discovered metabolic regulator. Interestingly, FGF21 is also known to inhibit Janus kinase 2 (JAK2)-signal transducer and activator of transcription 5 (STAT5) signaling from the GH receptor in the liver, where FGF21 mRNA is predominantly expressed. In this study, we tested the hypothesis that FGF21 gene expression in the liver is controlled by GH through STAT5. We found that GH injection to cattle increased FGF21 mRNA expression in the liver. Mapped by a 5'-rapid amplification of cDNA ends assay, transcription of the FGF21 gene in the bovine liver was mainly initiated from a nucleotide 24 bp downstream of a TATA box. The bovine FGF21 promoter contains three putative STAT5-binding sites. EMSA confirmed the ability of them to bind to liver STAT5 protein from GH-injected cattle. Chromatin immunoprecipitation assays demonstrated that GH administration increased the binding of STAT5 to the FGF21 promoter in the liver. Cotransfection analyses showed that GH induced reporter gene expression from the FGF21 promoter in a STAT5-dependent manner. GH also stimulated FGF21 mRNA expression in cultured mouse hepatocytes. These data together indicate that GH directly stimulates FGF21 gene transcription in the liver, at least in part, through STAT5. This finding, together with the fact that FGF21 inhibits GH-induced JAK2-STAT5 signaling in the liver, suggests a novel negative feedback loop that prevents excessive JAK2-STAT5 signaling from the GH receptor in the liver. (Endocrinology 153: 750-758, 2012)
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    Growth Hormone-Activated STAT5 May Indirectly Stimulate IGF-I Gene Transcription through HNF-3 gamma
    Eleswarapu, Satyanarayana; Ge, Xiaomei; Wang, Ying; Yu, Jie; Jiang, Honglin (Endocrine Society, 2009-12)
    IGF-I is abundantly expressed in the liver under the stimulation of GH. We showed previously that expression of hepatocyte nuclear factor (HNF)-3 gamma, a liver-enriched transcription factor, was strongly stimulated by GH in bovine liver. In this study, we determined whether GH-increased HNF-3 gamma 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-3 gamma 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-3 gamma 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-3 gamma 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-3 gamma 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-3 gamma 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-3 gamma gene. (Molecular Endocrinology 23: 2026-2037, 2009)
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    Roles of Growth Hormone, Insulin-Like Growth Factor I, and Sh3 and Cysteine Rich Domain 3 in Skeletal Muscle Growth
    Ge, Xiaomei (Virginia Tech, 2012-01-12)
    Three studies were conducted to achieve the following respective objectives: 1) to determine the cellular mechanism by which growth hormone (GH) stimulates skeletal muscle growth; 2) to identify the signaling pathways that mediate the different effects of insulin-like growth factor I (IGF-I) on skeletal muscle growth; and 3) to determine the role of a functionally unknown gene named SH3 and cysteine rich domain 3 (STAC3) in myogenesis. In the first study, the myogenic precursor cells, satellite cells, were isolated from cattle and allowed to proliferate as myoblasts or induced to fuse into myotubes in culture. GH increased protein synthesis without affecting protein degradation in myotubes; GH had no effect on proliferation of myoblasts; GH had no effect on IGF-I mRNA expression in either myoblasts or myotubes. These data suggest that GH stimulates skeletal muscle growth in cattle in part through stimulation of protein synthesis and that this stimulation is not mediated through increased IGF-I mRNA expression in the muscle. In the second study, the signaling pathways mediating the effects of IGF-I on proliferation of bovine myoblasts and protein synthesis and degradation in bovine myotubes were identified by adding to the culture medium rapamycin, LY294002, and PD98059, which are specific inhibitors of the signaling molecules mTOR, AKT, and ERK, respectively. The effectiveness of these inhibitors was confirmed by Western blotting. Proliferation of bovine myoblasts was stimulated by IGF-I, and this stimulation was partially blocked by PD98059 and completely blocked by rapamycin or LY294002. Protein degradation in myotubes was inhibited by IGF-I and this inhibition was completely relieved by LY294002, but not by rapamycin or PD98059. Protein synthesis in myotubes was increased by IGF-I, and this increase was completely blocked by rapamycin, LY294002, or PD98059. These data demonstrate that IGF-I stimulates proliferation of bovine myoblasts and protein synthesis in bovine myotubes through both the PI3K/AKT and the MAPK signaling pathways and that IGF-I inhibits protein degradation in bovine myotubes through the PI3K/AKT pathway only. In the third study, the potential roles of STAC3 in myoblast proliferation, differentiation, and fusion were investigated. Overexpression of STAC3 inhibited differentiation of C2C12 cells (a murine myoblast cell line) and fusion of these cells into myotubes, whereas knockdown of STAC3 had the opposite effects. Either STAC3 overexpression or STAC3 knockdown had no effect on proliferation of C2C12 cells. Myoblasts from STAC3-deficient mouse embryos had a greater ability to fuse into myotubes than control myoblasts; the former cells also expressed more mRNAs for the myogenic regulators MyoD and myogenin and the adult myosin heavy chain protein MyHC1 than the latter. These results suggest that STAC3 inhibits myoblast differentiation and fusion.
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    Stac3 Inhibits Myoblast Differentiation into Myotubes
    Ge, Xiaomei; Zhang, Yafei; Park, Sungwon; Cong, Xiaofei; Gerrard, David E.; Jiang, Honglin (PLOS, 2014-04-30)
    The functionally undefined Stac3 gene, predicted to encode a SH3 domain- and C1 domain-containing protein, was recently found to be specifically expressed in skeletal muscle and essential to normal skeletal muscle development and contraction. In this study we determined the potential role of Stac3 in myoblast proliferation and differentiation, two important steps of muscle development. Neither siRNA-mediated Stac3 knockdown nor plasmid-mediated Stac3 overexpression affected the proliferation of C2C12 myoblasts. Stac3 knockdown promoted the differentiation of C2C12 myoblasts into myotubes as evidenced by increased fusion index, increased number of nuclei per myotube, and increased mRNA and protein expression of myogenic markers including myogenin and myosin heavy chain. In contrast, Stac3 overexpression inhibited the differentiation of C2C12 myoblasts into myotubes as evidenced by decreased fusion index, decreased number of nuclei per myotube, and decreased mRNA and protein expression of myogenic markers. Compared to wild-type myoblasts, myoblasts from Stac3 knockout mouse embryos showed accelerated differentiation into myotubes in culture as evidenced by increased fusion index, increased number of nuclei per myotube, and increased mRNA expression of myogenic markers. Collectively, these data suggest an inhibitory role of endogenous Stac3 in myoblast differentiation. Myogenesis is a tightlycontrolled program; myofibers formed from prematurely differentiated myoblasts are dysfunctional. Thus, Stac3 may play a role in preventing precocious myoblast differentiation during skeletal muscle development.
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    Stac3 Is a Novel Regulator of Skeletal Muscle Development in Mice
    Reinholt, Brad M.; Ge, Xiaomei; Cong, Xiaofei; Gerrard, David E.; Jiang, Honglin (PLOS, 2013-04-23)
    The goal of this study was to identify novel factors that mediate skeletal muscle development or function. We began the study by searching the gene expression databases for genes that have no known functions but are preferentially expressed in skeletal muscle. This search led to the identification of the Src homology three (SH3) domain and cysteine rich (C1) domain 3 (Stac3) gene. We experimentally confirmed that Stac3 mRNA was predominantly expressed in skeletal muscle. We determined if Stac3 plays a role in skeletal muscle development or function by generating Stac3 knockout mice. All Stac3 homozygous mutant mice were found dead at birth, were never seen move, and had a curved body and dropping forelimbs. These mice had marked abnormalities in skeletal muscles throughout the body, including central location of myonuclei, decreased number but increased cross-sectional area of myofibers, decreased number and size of myofibrils, disarrayed myofibrils, and streaming Z-lines. These phenotypes demonstrate that the Stac3 gene plays a critical role in skeletal muscle development and function in mice.