Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.26.9 (ribonuclease)
6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Fetal growth is increased when pregnant gilts are treated with recombinant porcine somatotropin. The mechanism for increased fetal growth was examined by measuring the expression of IGF-I and -II and IGF-binding protein-2 (IGFBP-2) mRNA in liver and reproductive tissues of somatotropin- and saline-treated pregnant gilts. Twenty-four pregnant gilts received daily injections of either saline (control; n=12) or 5 mg recombinant porcine somatotropin (n=12) from day 30 to day 43 of gestation. Gilts were slaughtered on day 44 of gestation and liver, ovary, placenta, placental uterus (uterus with adjacent placental tissue) and non-placental uterus (region of the necrotic tip) were collected. The mRNAs for somatotropin receptor, IGFs -I and -II, IGFBP-2 and pregnancy-associated glycoprotein (a marker of trophoblast tissue) were analyzed by Northern blotting or ribonuclease protection assay. Gilts treated with somatotropin had heavier fetuses and placentas. The concentration of mRNA for the components of the IGF system was tissue-dependent. The uterine IGF-I mRNA concentration was greater in non-placental than in placental uterus. The greatest IGF-II mRNA concentration was observed in placenta, and adjacent uterine tissue expressed IGFBP-2 mRNA intensely. In non-placental uterus, IGFBP-2 mRNA was nearly undetectable. Somatotropin-dependent regulation of IGF-I was only observed in liver, where the greatest somatotropin receptor mRNA concentration was found. In the pregnant uterus, somatotropin failed to change the concentration of IGF or IGFBP-2 mRNA. Pregnancy-associated glycoprotein mRNA concentration was decreased by somatotropin. In summary, increased fetal growth in somatotropin-treated pregnant pigs was not associated with changes in IGF or IGFBP-2 mRNA concentration in reproductive tissues. Other mechanisms, therefore, lead to enhanced fetal growth in somatotropin-treated pregnant pigs.
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PMID:Insulin-like growth factor (IGF)-I, IGF-II, IGF-binding protein-2 and pregnancy-associated glycoprotein mRNA in pigs with somatotropin-enhanced fetal growth. 983 61

In the mouse, GH-binding protein (GHBP) and GH receptor (GHR) are encoded by a single gene via alternative splicing. We previously demonstrated that the steady-state levels of the GHR and GHBP mRNAs are significantly elevated in mouse liver during pregnancy. Hepatic GHR and GHBP mRNAs are associated primarily with one of two different 5' untranslated regions (5' UTRs), designated 5' UTR Liver1 (L1) and Liver2 (L2). Distinct promoters associated with each of these 5' UTRs have recently been characterized. In the present study, we have investigated the role of transcriptional activation in the pregnancy-induced upregulation of GHR and GHBP mRNAs in liver. We also report on the relative contribution of the 5' UTR L1 and 5' UTR L2 promoters to the hepatic expression of the GHR/GHBP gene in the liver. Our approach was to compare, by ribonuclease protection assay (RPA), GHR/GHBP transcript levels in hepatic nuclear and total cellular RNA samples from virgin and late-pregnant mice. In these RPAs we utilized riboprobes that were complementary to the coding region of GHR/GHBP transcripts, as well as to the two noncoding, alternative first exons 5' UTR L1 and L2. When employing the coding region probe, RPAs revealed that the gestational increase in the levels of nuclear GHR/GHBP transcripts were statistically comparable with the increase in GHR/GHBP transcript levels in total cellular RNA. This finding suggests that enhanced transcriptional activity, rather than increased cytoplasmic half-life, is responsible for the upregulation of GHR/GHBP RNA in the pregnant liver. In RPAs utilizing the noncoding region probes, both nuclear and total cellular GHR/GHBP transcripts associated with 5' UTR L1 were significantly upregulated in late-pregnant as compared with virgin mice. In contrast, the levels of both nuclear and total GHR/GHBP transcripts associated with 5' UTR L2 were comparable between nonpregnant and pregnant animals. Moreover, 5' UTR L2-containing transcripts were present at levels that were only 3-5% of the 5' UTR L1-associated transcripts in the late-pregnant liver. Thus, we conclude that the gestational upregulation of the GHR/GHBP gene in the mouse liver can be ascribed to the significantly enhanced transcriptional activity of the 5' UTR L1 promoter.
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PMID:Transcriptional upregulation of hepatic GH receptor and GH-binding protein expression during pregnancy in the mouse. 1042 50

The growth hormone (GH) receptor is essential for the actions of growth hormone on postnatal growth and metabolism. GH receptor transcripts are characterized by the presence of disparate 5'-untranslated exons. Factors regulating the expression of the GC rich L2 transcript of the murine GH receptor gene have hitherto remained unidentified. To characterize the mechanisms regulating expression of the L2 transcript, primer extension and ribonuclease protection assays were used to identify transcription start sites in RNA from liver of adult mice. Transient transfection experiments revealed that 2.0 kilobase pairs of the L2 5'-flanking sequence exhibited promoter activity in BNL CL.2 (mouse liver) cells, CV-1 (monkey kidney) cells, and HRP.1 trophoblasts. Deletional analysis localized a major regulatory region to within 75 base pairs of the 5' transcription start site. Sequence analysis revealed that the region contained consensus binding sites for the Sp family of transcription factors. Standard gel shift and supershift analysis using liver nuclear extracts established that Sp1 and Sp3 bound this regulatory element. Transfection of wild type but not mutant decoy oligonucleotides into BNL CL.2 cells decreased the activity of the L2 promoter. Overexpression of Sp1 and Sp3 protein in Drosophila Schneider cells established that Sp3 is more potent than Sp1 in transactivating the L2 promoter. Co-transfection experiments further established that Sp1 antagonizes the activity of Sp3 to transactivate the L2 promoter. Western blot analysis of liver nuclear extracts revealed that the levels of Sp3 increase significantly after birth, suggesting a role for the Sp family of transcription factors in controlling the fetal to postnatal increase in GH receptor gene expression.
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PMID:Role of the Sp family of transcription factors in the ontogeny of growth hormone receptor gene expression. 1056 9

The growth hormone (GH) receptor gene is characterized by heterogeneity in the 5'-untranslated region (UTR). The technique of 5'-rapid amplification of cDNA ends (RACE) was employed to identify potentially novel 5'-UTRs for the GH receptor gene. One of the RACE clones displayed sequence homology to the human V5-UTR; hence this transcript was designated as L5. Sequence analysis of genomic DNA established that L5 was immediately upstream of exon 2. Northern blot analysis indicated that two bands of sizes congruent with4.8 kb, corresponding to GH receptor mRNA, and congruent with1.5 kb corresponding to GH binding protein mRNA, were detectable in liver, skeletal muscle, kidney and heart but not in brain, spleen, lung or testis. Fluorescent 5'-nuclease real-time RT-PCR based analysis indicated that in the placenta and fetal liver, the L5 transcript represented 10-15% of the GH receptor transcripts. In the adult liver, heart and kidney, the L5 transcript is less abundant accounting for 1-5% of the total GH receptor transcripts. Primer extension and ribonuclease protection assays were performed to identify the major transcription start site at 778 bp from the ATG codon. Transient transfection experiments revealed that the 5'-flanking sequence had promoter activity in rat placental trophoblast (HRP.1), Chinese hamster ovary (CHO) and mouse liver (BNL CL.2) cells. Analysis of expression of the L5 transcript in the non-obese diabetic (NOD) mouse, a model of spontaneous autoimmune diabetes, indicated that the expression of the L5 transcript was decreased in liver and kidney by 80-90 and 40-50%, respectively, but expression remained unchanged in the heart.
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PMID:Identification and characterization of a novel transcript of the murine growth hormone receptor gene exhibiting development- and tissue-specific expression. 1116 47

Elevation of circulating GH acts to feed back at the level of the hypothalamus to decrease GH-releasing hormone (GHRH) and increase somatostatin (SRIF) production. In the rat, GH-induced changes in GHRH and SRIF expression are associated with changes in pituitary GHRH receptor (GHRH-R), GH secretagogue receptor (GHS-R), and SRIF receptor subtype messenger RNA (mRNA) levels. These observations suggest that GH regulates its own synthesis and release not only by altering expression of key hypothalamic neuropeptides but also by modulating the sensitivity of the pituitary to hypothalamic input, by regulating pituitary receptor synthesis. To further explore this possibility, we examined the relationship between the expression of hypothalamic neuropeptides [GHRH, SRIF, and neuropeptide Y (NPY)] and pituitary receptors [GHRH-R, GHS-R, and SRIF receptor subtypes (sst2 and sst5)] in two mouse strains with alterations in the GH-axis; the GH receptor/binding protein gene-disrupted mouse (GHR/BP-/-) and the metallothionein promoter driven human GHRH (MT-hGHRH) transgenic mouse. In GHR/BP-/- mice, serum insulin-like growth factor I levels are low, and circulating GH is elevated because of the lack of GH negative feedback. Hypothalamic GHRH mRNA levels in GHR/BP-/- mice were 232 +/- 20% of GHR/BP+/+ littermates (P < 0.01), whereas SRIF and NPY mRNA levels were reduced to 86 +/- 2% and 52 +/- 3% of controls, respectively (P < 0.05; ribonuclease protection assay). Pituitary GHRH-R and GHS-R mRNA levels of GHR/BP-/- mice were elevated to 275 +/- 55% and 319 +/- 68% of GHR/BP+/+ values (P < 0.05, respectively), whereas the sst2 and sst5 mRNA levels did not differ from GHR/BP intact controls as determined by multiplex RT-PCR. Therefore, in the absence of GH negative feedback, both hypothalamic and pituitary expression is altered to favor stimulation of GH synthesis and release. In MT-hGHRH mice, ectopic hGHRH transgene expression elevates circulating GH and insulin-like growth factor I. In this model of GH excess, endogenous (mouse) hypothalamic GHRH mRNA levels were reduced to 69 +/- 6% of nontransgenic controls, whereas SRIF mRNA levels were increased to 128 +/- 6% (P < 0.01). NPY mRNA levels were not significantly affected by hGHRH transgene expression. Also, MT-hGHRH pituitary GHRH-R and GHS-R mRNA levels did not differ from controls. However, sst2 and sst5 mRNA levels in MT-hGHRH mice were increased to 147 +/- 18% and 143 +/- 16% of normal values, respectively (P < 0.05). Therefore, in the presence of GH negative feedback, both hypothalamic and pituitary expression is altered to favor suppression of GH synthesis and release.
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PMID:The growth hormone (GH)-axis of GH receptor/binding protein gene-disrupted and metallothionein-human GH-releasing hormone transgenic mice: hypothalamic neuropeptide and pituitary receptor expression in the absence and presence of GH feedback. 1118 26

The signal transduction pathways that mediate GH-dependent regulation of IGF-I gene expression remain poorly defined. To establish a GH-responsive in vitro model system to study the effect of GH on the expression of the endogenous IGF-I gene, primary hepatocytes from adult male rats were prepared. These cells expressed both the GH receptor and the IGF-I gene, as demonstrated using a ribonuclease protection assay. Western blot analyses using antibodies directed against the phosphorylated forms of the ERKs, signal transducer and activator of transcription-5, and Akt/protein kinase B, a protein kinase that is downstream of PI3K, demonstrated GH-dependent phosphorylation of these signaling molecules. These signaling molecules are components of the major signal transduction pathways that are activated by GH. To determine whether GH-responsive IGF-I gene expression could be demonstrated in these cells, hepatocytes were treated without or with 50 ng/ml GH for 3--48 h. IGF-I mRNA levels increased within 3 h, and a maximal 2.2-fold increase was observed after 24 h of GH treatment. To determine whether ERK activation contributes to GH-induced IGF-I gene expression, hepatocytes were treated for 12 h without or with 50 ng/ml GH and 50 microM PD98059, an inhibitor of MAPK kinase-1 and -2. Treatment with PD98059 did not have a significant effect on GH-induced IGF-I gene expression. Similar studies were performed using 50 microM LY 294002, an inhibitor of PI3K. These studies demonstrated that treatment with LY 294002 completely abrogated GH-induced IGF-I gene expression. In contrast, PI3K-specific doses of another inhibitor of PI3K, wortmannin, failed to inhibit the GH-induced increase in IGF-I mRNA levels. In summary, rat hepatocytes in primary culture provide a good model system to study GH-induced IGF-I gene expression, and the results of these studies suggest that a signaling pathway inhibited by LY 294002, possibly a PI3K-dependent pathway, is important for GH-stimulated IGF-I gene expression in hepatocytes.
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PMID:LY 294002, an inhibitor of phosphatidylinositol 3-kinase, inhibits GH-mediated expression of the IGF-I gene in rat hepatocytes. 1151 77

We investigated the effect of increasing nutrient intake on the responsiveness of the GH/IGF-I system in calves fed a high-protein milk replacer. Fifty-four Holstein bull calves were fed one of three levels (low, medium, and high; n = 18 per treatment) of a 30% crude protein, 20% fat milk replacer to achieve target rates of gain of 0.50, 0.95, or 1.40 kg/d, respectively, for low, medium, and high. Six calves per treatment were slaughtered at approximately 65, 85, and 105 kg BW. Additionally, six calves were slaughtered at 1 d of age to provide baseline data. Plasma aliquots from blood samples collected weekly were analyzed for IGF-I, insulin, glucose, NEFA, and plasma urea nitrogen (PUN). Plasma IGF-I and insulin, measured weekly, increased (P < 0.001) with greater nutrient intake from wk 2 of life to slaughter. Plasma glucose and NEFA also increased (P < 0.05) with nutrient intake. In addition, each calf underwent a GH challenge beginning 4 d before the scheduled slaughter. Plasma from blood collected before the first GH injection and 14 and 24 h after the third injection was analyzed for IGF-I and PUN. Response to challenge, calculated as the absolute difference between the prechallenge and 14-h postchallenge plasma IGF-I concentrations, was significant in calves on all three treatments. Plasma urea nitrogen was not different among treatments as measured weekly but decreased (P < 0.001) following GH challenge in all calves. Results of ribonuclease protection assays showed increased expression of hepatic mRNA for GH receptor 1A and IGF-I with increased intake. The amounts of GH receptor and IGF-I mRNA in muscle and adipose, however, were not affected by intake. In summary, plasma IGF-I was elevated in calves with increased nutrient intake, and the elevations in plasma IGF-I following short-term administration of GH were significant in all calves by 65 kg BW. Data demonstrate that in well-managed milk-fed calves the somatotropic (GH/IGF-I) axis is functionally coordinated and sensitive to nutrient intake and GH.
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PMID:Effect of nutrient intake on the development of the somatotropic axis and its responsiveness to GH in Holstein bull calves. 1207 34


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