Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The biological activity of insulin-like growth factor-I (IGF-I) is mediated by a transmembrane glycoprotein (type-1 IGF receptor or IGF-I receptor) that shows considerable sequence homology with the insulin receptor. In order to detect the expression of this gene in chicken liver tissue, a plasmid was constructed containing a fragment of chicken IGF-I receptor cDNA. The cDNA fragment corresponded to nucleotides 326-599 of the human IGF-I receptor cDNA and showed 86.1 and 69.3% homology at the nucleotide level and 96.7 and 80.2% homology at the amino acid level with the human IGF-I receptor and insulin receptor respectively. The construct was used to generate an antisense RNA probe for the detection of IGF-I receptor mRNA transcripts in 1- and 4-week-old chick liver tissue. IGF-I receptor gene expression was initially detected by the reverse transcriptase polymerase chain reaction using synthetic chicken IGF-I receptor oligonucleotides. Amplified fragments of the correct size were detected in both RNA samples. Northern blots were also used to detect IGF-I receptor mRNA transcripts in the liver RNA samples. The results indicated that the amount of receptor mRNA decreased significantly between 1 and 4 weeks after hatch. In contrast, chicken beta-actin gene expression remained constant over this period. A major IGF-I receptor RNA transcript (11 kb) was observed in blots from 1-week-old livers, less abundant transcripts were also observed ranging in size from 8 to 9 kb.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The expression of a putative insulin-like growth factor-I receptor gene in the liver of the developing chick. 132 34

Currently available evidence supports the hypothesis that insulin-like growth factor-I (IGF-I) secreted by small preantral follicles may be involved in stimulating the initial differentiation of the theca interna and, in particular, expression of the LH receptor in pre-theca cells. To test this hypothesis, we examined the effects of IGF-I on LH receptor mRNA expression in theca-interstitial cells (TIC) isolated from the ovaries of hypophysectomized immature rats by percoll gradient centrifugation. TIC (3.5 x 10(4) viable cells/well) were cultured up to 6 days with and without LH (0-10 ng/ml) and IGF-I (0-100 ng/ml). Androsterone in the medium was measured by RIA, and LH receptor mRNA was measured by specific reverse transcriptase-polymerase chain reaction assay. LH receptor mRNA was low in control (untreated) TIC. IGF-I stimulated a dose-related increase (2-fold) in LH receptor mRNA at 2 days (ED50 = 9.0 +/- 1.9 ng/ml) that remained constant at 4 days and then declined to basal levels at 6 days. LH stimulated a dose-related (ED50 = 17.6 +/- 1.0 pg/ml) increase in LH receptor mRNA that reached a maximum of 4-fold at 2 days. At 4 days, LH down-regulated LH receptor mRNA below basal levels, and it had no effect at 6 days. Addition of IGF-I (30 ng/ml) to LH-treated TIC abolished the stimulatory effect of LH throughout the culture period. LH receptor mRNA was highly sensitive to LH since the ED50 was approximately 2.5-fold lower than for stimulation of androsterone production (39.8 +/- 3.8 pg/ml). To understand the molecular mechanism of the synergistic stimulation of androgen production by IGF-I and LH, the effects of IGF-I on the cAMP/protein kinase A (PKA) signaling pathway were examined. When freshly isolated TIC were challenged with IGF-I alone (30 ng/ml), there was no effect on cAMP production or PKA activity, but IGF-I augmented LH stimulation of cAMP production slightly at high concentrations of LH and blocked stimulation of PKA activity by a saturating concentration of LH (3 ng/ml), suggesting that IGF-I increased LH down-regulation of PKA. We next examined the effects of IGF-I on LH receptor number. When TIC were placed into culture, LH/hCG binding sites decreased to approximately 35% of the initial number at 24 h and 25% at 2 days. This decrease was accompanied by a similar loss of cholera toxin- and hCG-stimulated cAMP production.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Insulin-like growth factor-I regulation of luteinizing hormone (LH) receptor messenger ribonucleic acid expression and LH-stimulated signal transduction in rat ovarian theca-interstitial cells. 752 76

An alternatively spliced transcript of the human insulin-like growth factor-I (IGF-I) gene is described. The transcript was identified in human liver RNA by reverse transcriptase-polymerase chain reaction, cloning, and sequencing. It contained IGF-I exons 3 and 4, 49 basepairs of exon 5, then exon 6 (exon 4-5-6). The 5'-donor site at the exon 5-6 junction was a cryptic 5'-donor splice site (IGF633). The 3'-acceptor site of the splice was the usual intron-exon 6 junction. A second pair of primers across the exon 5-exon 6 junction was used to confirm the presence of the transcript by reverse transcriptase-polymerase chain reaction. Cloning and sequencing this second fragment confirmed the presence of this splice in human liver. The exon 4-5-6 transcript was quantified at about 10% relative to the exon 4-6 transcript in human livers (n = 7 subjects), but was not detected in other tissues. The exon 4-5-6 transcript was found in cultured human hepatoma HepG2 cells and increased, relative to exon 4-6 transcripts, in response to GH, but not in cultured human lymphoblast IM-9 cells. The exon 4-5-6 splice predicts a prepro-IGF-I of 158 amino acid residues, with an E-peptide sequence of 24 residues (Ec). The deduced Ec peptide sequence is 73% homologous to the rat Eb-peptide sequence. The predicted final residues of the Ec peptide are frameshifted exon 6 codons ending in an in-frame stop codon. The predicted peptide sequences of Ec and Eb differ at the cleavage site of the Eb-peptide fragment (IBE1), which has been shown to have mitogenic activity. These data suggest that 1) the exon 4-5-6 splice has hepatic tissue expression and occurs by the use of a cryptic 5'-donor consensus splice site (IGF633) in exon 5; 2) exon 4-5-6 can be hormonally regulated in cultured human HepG2 cells; 3) exon 4-5-6 is the human counterpart of the rat IGF-IEb, because the complementary DNA and predicted sequences are homologous; and 4) the production of IBE1 is potentially regulated by alternative splicing.
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PMID:An alternatively spliced human insulin-like growth factor-I transcript with hepatic tissue expression that diverts away from the mitogenic IBE1 peptide. 772 Jun 41

We cloned four kinds of goat insulin-like growth factor-I (IGF-I) cDNAs. Each of them encodes the same mature IGF-I and different signal peptides. In this study, we analyzed expression of the four kinds of mRNAs in tissues in various developmental stages by a reverse transcriptase-polymerase chain reaction assay. The results suggest that their expression may be controlled by different promoters, the activities of which are tissue- and development-specific.
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PMID:Tissue- and development-specific expression of goat insulin-like growth factor-I (IGF-I) mRNAs. 777 48

Insulin-like growth factor-I (IGF-I) is expressed not only in liver, but also in brain and other tissues. This ubiquitous IGF-I has a complex pattern of expression due to multiple transcription start sites, polyadenylation sites and exon skipping. We have isolated a cDNA encoding a brain-specific IGF-I from a catfish brain cDNA library. Also, a fragment encoding ubiquitous IGF-I was amplified from brain and liver mRNA and the deduced protein shown to be distinct (66% sequence identity) from brain-specific IGF-I. Consistent with other IGF-I prepropeptides, the brain-specific IGF-I has a 43-residue signal peptide followed by B, C, A, D, and E domains. Retained in the catfish brain-specific IGF-I peptide are residues predicted to be involved with the correct tertiary folding, disulfide linkages, and receptor binding. Northern blot analysis of poly(A+)-rich mRNA from brain indicated a single 1600-base pair transcript; a band was not detected from mRNA of liver, stomach, pancreas, pituitary, blood, herring brain, or brain poly(A-) RNA. A sensitive reverse transcriptase/polymerase chain reaction assay also showed that brain-specific IGF-I mRNA was expressed solely in the Thai catfish brain but not liver, stomach, pancreas, pituitary, ovary, and African catfish brain.
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PMID:Catfish express two forms of insulin-like growth factor-I (IGF-I) in the brain. Ubiquitous IGF-I and brain-specific IGF-I. 791 63

Previous studies using cultures of fetal rodents calvaria have indicated an important role for insulin-like growth factor-I (IGF-I) in the local control of bone formation. In this study, we have examined the expression of IGF-I in adult human osteoblast-like (hOB) cells. To detect very low levels and to distinguish between the two IGF-I mRNA transcripts Ea and Eb, which are formed by alternate splicing, we have used the reverse transcriptase-polymerase chain reaction (RT-PCR). Expression of both Ea and Eb IGF-I mRNA transcripts were found in human liver and in placenta. However, neither Ea nor Eb IGF-I mRNA could be detected under basal conditions or after stimulation with growth hormone in normal hOB cells and two human osteosarcoma cell lines with osteoblastic properties, SaOS-2 and MG-63. We conclude that adult hOB cells do not synthesize IGF-I. Thus, in contrast to its crucial role as a local regulator of skeletal remodeling in fetal rodent bone, IGF-I does not appear to have this autocrine function in adult human bone.
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PMID:Adult human osteoblast-like cells do not express insulin-like growth factor-I. 812 49

The insulin receptor possesses tyrosine kinase activity which is thought to mediate the biological effects of insulin upon target cells. pp120 is a liver-specific glycoprotein of apparent molecular size of 120 kDa that is phosphorylated on tyrosine residues by the receptors for insulin, insulin-like growth factor-I, and epidermal growth factor. Previously, we demonstrated that pp120 is identical to a liver-specific ecto-ATPase. In the present study, we have cloned the rat gene encoding pp120/ecto-ATPase. The gene is contained within approximately 15 kilobases of genomic DNA, and consists of nine exons interrupted by eight introns. Using the reverse transcriptase/polymerase chain reaction, we isolated cDNA clones complementary to rat liver mRNA encoding pp120/ecto-ATPase. Sequence analysis indicated the presence of two populations of cDNA's that differ by the presence or absence of a 53-base pair (bp) fragment encoding the juxta-membrane region of the cytoplasmic domain. By cloning the corresponding region of the ecto-ATPase gene, we demonstrated that the 53-bp represents exon 7 of the gene. This 53-bp exon undergoes alternative splicing, thereby giving rise to two mRNA variants. Deletion of this 53-bp cassette exon introduces a frameshift, and results in a premature chain termination codon that truncates the cytoplasmic domain. The truncated cytoplasmic domain contains 10 rather than 71 amino acid residues. Because the short isoform of ecto-ATPase lacks the putative sites for tyrosine- and serine-specific phosphorylation, this alternative splicing may have a major effect upon the physiological function of the enzyme.
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PMID:pp120/ecto-ATPase, an endogenous substrate of the insulin receptor tyrosine kinase, is expressed as two variably spliced isoforms. 838 Apr 6

Alternative splicing of ovine insulin-like growth factor-I (IGF-I) transcripts generates three different mRNAs. Class 1 and class 2 transcripts contain exons 1 and 2 spliced to exon 3, respectively. A novel IGF-I mRNA containing exon W is spliced to exon 3 and has been located upstream of exon 1. No in-frame methionine codon was present in exon W and therefore translation is proposed to initiate at the methionine codon present in exon 3. Using primer extension, transcription initiation sites were found 179, 336, and 368 nucleotides upstream of exon 1 and 86, 96, 131, and approximately 850 nucleotides upstream of exon 2. The locations of these transcription initiation sites are well conserved among mammalian and avian IGF-I genes. Expression of exon 1-, 2-, and W-specific transcripts was examined in brain, heart, kidney, liver, lung, skeletal muscle, and spleen from adult ewes or 75-day fetal lambs using a reverse transcriptase-polymerase chain reaction assay. Exon 1 transcripts were the most abundant and found in all fetal and adult tissues. Exon 2 transcripts were found in all tissues and were generally expressed the highest in adult liver. Exon W transcripts were also found to be expressed in all tissues examined. Thus, the three alternatively spliced ovine IGF-I transcripts were expressed in a variety of fetal and adult tissues.
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PMID:Characterization of multiple transcription initiation sites of the ovine insulin-like growth factor-I gene and expression profiles of three alternatively spliced transcripts. 846 47

Leukocytes synthesize a variety of hormones that were once thought to be unique products of endocrine tissues. Understanding the regulation of leukocyte-derived hormone synthesis requires an accurate means for measuring steady-state expression of specific mRNA transcripts. Here we describe a competitive reverse transcriptase-polymerase chain reaction (RT-PCR) technique to accurately quantitate macrophage-derived insulin-like growth factor-I (IGF-I) mRNA, and demonstrate the utility of this approach for measuring expression of leukocyte-derived hormone transcripts. A riboprobe was constructed to generate approximately 1 kb of synthetic competitor IGF-I RNA (exons 1 and 3-6) that differed from cellular IGF-I RNA by insertion of 122 bp of beta-actin RNA. One set of oligonucleotide primers could thus be used to simultaneously reverse transcribe and amplify both 144 bp of cellular (exons 3 and 4) and 266 bp of competitor IGF-I RNA. Densitometric scanning of the PAGE-separated PCR products revealed that the ratio of competitor to cellular amplified DNA bore a linear relationship (r2 > or = 0.98) to the amount of competitor RNA for both rat liver and splenocytes. However, rat liver contained 104 x 10(6) IGF-I molecules per microgram of total cellular RNA compared to only 2 x 10(6) IGF-I molecules for splenocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Competitive reverse transcriptase-polymerase chain reaction using a synthetic internal RNA standard to quantitate transcripts for leukocyte-derived hormones. 852 83

In mouse preadipocyte Ob1771 cells, transcription of the insulin-like growth factor-I (IGF-I) gene was stimulated by growth hormone (GH), and IGF-I protein combined with GH in medium was required for their differentiation to adipocytes. During induction of the differentiation, the intracellular expression of each class of IGF-I mRNA was analyzed by reverse transcriptase-polymerase chain reaction. When the cells were cultured in the presence of GH, the class 1del. IGF-I mRNA was a major molecular species among IGF-I mRNAs. In the presence of both GH and IGF-I, the splicing pattern of IGF-I mRNA changed from class 1del. to class 1. Moreover, as detected by Western blotting, the IGF-I protein was present in cells and in the medium only when the cells were cultured in the presence of both GH and IGF-I. We found that IGF-I secreted from Ob1771 cells could act in an autocrine/paracrine fashion and induce the differentiation of other Ob1771 cells. It was demonstrated that the translation efficiency of class 1 mRNA was higher than that of class 1del. mRNA in vitro. These results suggested that stimulation with exogenous IGF-I in the presence of GH was required for the production of class 1 IGF-I mRNA and that the production of the IGF-I protein was activated by increasing the translation efficiency through shifting the splicing pattern of IGF-I mRNA from class 1del. to class 1. Exogenous IGF-I triggered the differentiation by initiating the synthesis of endogenous IGF-I.
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PMID:Regulation of insulin-like growth factor-I expression in mouse preadipocyte Ob1771 cells. 862 20


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