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:3.1.27.1 (RNase)
16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Restriction of dietary protein consumption of young male rats results in decreased growth velocity and a reduction in the abundance of hepatic IGF-I mRNA. It is not known whether the reduction of IGF-I mRNA abundance in the liver of protein-restricted rats results from a decrease in IGF-I gene transcription. In the present study, three experiments were performed with 4-week-old male rats to examine the effect of protein restriction on IGF-I gene transcription in liver. In these experiments, we monitored IGF-I nuclear transcripts (pre-mRNA) within total cellular RNA using a ribonuclease protection assay. In the first experiment, a consistent decrease in IGF-I mRNA from animals fed isocaloric diets containing 20% (control), 12%, 8% and 4% protein (dietary effect, P < 0.001) was not paralleled by a decrease (P > 0.50) in IGF-I pre-mRNA. Two additional experiments examining the effect of 4% vs 20% protein diets yielded comparable results. Pooled results from these two studies (n = 12/treatment) demonstrated that a 64% reduction (P < 0.0001) in IGF-I mRNA abundance was not accompanied by a decrease in IGF-I pre-mRNA (1.17 vs 1.31 +/- 0.21 image density units for 4% and 20% protein treatments). Unlike IGF-I, the abundance of carbamyl phosphate synthetase-I (CPS-I) pre-mRNA and mRNA was comparably reduced (approximately 70%, P < 0.001), indicating that the decrease in mRNA of this urea cycle enzyme during protein restriction occurs predominantly by a transcriptional mechanism. A common feature of all experiments was a pronounced variability in the expression of hepatic IGF-I pre-mRNA among animals, which was not diet specific. To test whether the variability in IGF-I gene transcription was correlated with variability in the transcription of another gene that is regulated by GH, we quantified the abundance of nuclear transcripts for the serine protease inhibitor 2.1 (SPI 2.1) gene. A positive association (r = 0.81, P < 0.0001) between SPI 2.1 and IGF-I nuclear transcripts was demonstrated. The correlation between IGF-I and SPI 2.1 transcripts was specific, because the quantity of IGF-I and CPS-I nuclear transcripts was not correlated in this study. Although transcription of the IGF-I and SPI 2.1 genes was similar, the abundance of SPI 2.1 mRNA was not altered by protein deprivation.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:IGF-I and serine protease inhibitor 2.1 nuclear transcript abundance in rat liver during protein restriction. 763 24

Glucocorticoids have a number of effects on bone cell function, some of which might be mediated by changes in the synthesis or activity of insulin-like growth factors (IGFs). Glucocorticoids inhibit IGF-I, but not IGF-II, synthesis in osteoblasts and decrease the expression of selected IGF-binding proteins. The effects of glucocorticoids on IGF-I and -II receptor messenger RNA (mRNA) expression in osteoblasts are not known, and changes in IGF-I or -II receptor levels could result in changes in IGF activity. We examined the effects of glucocorticoids on IGF-I and -II receptor mRNA expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cells). Cortisol at 1 microM for 2-48 h did not alter IGF-I receptor transcripts, as determined by Northern blot analysis and ribonuclease protection assay. In contrast, cortisol caused a time- and dose-dependent inhibition of IGF-II receptor mRNA levels. The effect was maximal at 0.1-1 microM for 24-48 h and was accompanied by a decrease in IGF-II receptor levels, as determined by affinity labeling, cross-linking and polyacrylamide gel electrophoresis, Western immunoblot, and Scatchard analysis. The effect of cortisol on IGF-II receptor transcripts was not dependent on de novo protein synthesis. Cortisol did not modify the IGF-II receptor mRNA half-life in transcriptionally arrested Ob cells and decreased the rate of IGF-II receptor RNA transcription in nuclear run-on assays. In conclusion, cortisol decreases transcription of the IGF-II receptor in Ob cell cultures, an effect that could mediate selected actions of glucocorticoids in bone.
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PMID:Cortisol represses insulin-like growth factor II receptor transcription in skeletal cell cultures. 766 43

Abnormalities of GH secretion and clearance are well-documented in poorly controlled insulin-dependent diabetes mellitus (IDDM), but the contribution of the receptor (GHR) and the GH-binding protein (GHBP) to these abnormalities has not been defined. We studied the expression of the GHR/GHBP gene in the livers, hearts and kidneys in streptozocin-induced diabetes (STZ-D) in the rat. GHR and GHBP mRNA levels were measured by Northern blot and ribonuclease protection assays. Whereas levels of GHR and GHBP mRNA were significantly decreased in liver and heart of STZ-D rats when compared with the control group (P < 0.01), GHR mRNA was significantly increased in the kidneys of STZ-D rats (P = 0.03). Six days of insulin treatment did not significantly alter the levels of GHR/GHBP mRNA in the liver or heart of STZ-D rats, but significantly decreased GHBP mRNA (P = 0.04) in the kidney. Circulating IGF-I was reduced, as was IGF-I mRNA in the liver and heart of STZ-D rats; only circulating IGF-I was restored by insulin treatment. Neither STZ-D nor insulin treatment affected IGF-I or IGF-I receptor mRNA concentrations in the kidney. We conclude that (1) STZ-D modulates the expression of the GHR/GHBP gene and (2) that these changes in GHR/GHBP mRNA concentrations are tissue-specific; STZ-D decreases GHR/GHBP mRNA in liver and heart tissue but increases GHR mRNA concentrations in the kidney. Our results indicate a role for decreased numbers of hepatic GHRs in the pathogenesis of resistance to GH's actions in terms of IGF-I generation and promotion of linear growth in IDDM. We postulate that increased GHR expression in the kidney may be involved in the renal complications of IDDM.
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PMID:Tissue-specific regulation of the growth hormone receptor gene in streptozocin-induced diabetes in the rat. 796 96

Salmon have been shown to express alternatively spliced IGF-I mRNA transcripts coding for four different IGF-I prohormones. These transcripts, now designated Ea-1, Ea-2, Ea-3 and Ea-4, differ in size due to the inclusion of additional sequences in the E domain-coding region of the molecule. In this study, the tissue distribution and hormonal regulation of expression of alternatively spliced IGF-I mRNA transcripts were investigated in coho salmon. IGF-I mRNAs were detected by solution hybridization/RNase protection assay in all tissues examined. GH treatment significantly increased hepatic IGF-I mRNA content. Hepatic IGF-I mRNA levels were not influenced by prolactin or somatolactin. Heart, fat, brain, kidney, spleen and ovary IGF-I mRNA levels were not affected by GH, prolactin or somatolactin. Ea-1, Ea-3 and Ea-4 mRNA transcripts were detectable in the liver, and Ea-1 and Ea-3 levels increased dramatically in response to GH treatment, whereas the amount of Ea-4 mRNA was unchanged. Most non-hepatic tissues expressed only the Ea-4 transcript, and expression was not influenced by GH, prolactin or somatolactin. Ea-1 and Ea-3 transcripts were visible in gill samples from fish treated with GH. The ovaries of juvenile fish expressed Ea-1, Ea-2 and Ea-4. The amounts of these transcripts were not changed by gonadotrophin treatment. During smoltification of juvenile coho salmon, liver and gill IGF-I mRNA levels increased with increasing plasma GH and thyroxine concentrations. Muscle, brain and ovary IGF-I mRNA levels were unchanged during this period. These data suggest that the liver is a major site of IGF-I production in response to GH. Heart, fat, brain, kidney, spleen and ovary did not show increased IGF-I mRNA levels in response to GH treatment. GH and prolactin had inconsistent effects on muscle IGF-I mRNA levels. Somatolactin and a gonadotrophin preparation did not stimulate IGF-I expression in tissues of juvenile fish. Differences in tissue GH responsiveness can be partially explained by the expression of alternatively spliced IGF-I mRNAs. Of the four hepatic IGF-I mRNA transcripts, Ea-1 and Ea-3 are GH-responsive, while Ea-2 and Ea-4 are not. Most non-hepatic tissues express only the Ea-4 transcript, and IGF-I mRNA levels do not increase after GH treatment. The increased IGF-I mRNA levels observed in gill tissue during smoltification suggest that other factors, in addition to GH, may regulate IGF-I expression.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Differential expression and hormonal regulation of alternatively spliced IGF-I mRNA transcripts in salmon. 818 11

Two distinct class 1 and class 2 rat liver IGF-I mRNAs contain different 5' leader exons, 1 and 2. RNase protection, primer extension, RACE PCR and ribonuclease H mapping established the complete structure of the 5' end of class 1 and class 2 IGF-I mRNAs. Two major transcription start sites in exon 1 yield class 1 IGF-I mRNAs, including 345 or 245 bases of exon 1. Multiple, clustered transcription start sites in exon 2 yield class 2 IGF-I mRNAs with 84-50 bases of exon 2. Cell-free translation of in vitro transcribed IGF-I mRNAs suggests that class 1 and class 2 mRNAs preferentially initiate translation at distinct AUG codons to result in IGF-I precursors with either 48 residue class 1 pre-peptides or 32 residue class 2 pre-peptides. Some translation initiation also occurs at a downstream AUG common to class 1 and 2 mRNAs to yield IGF-I precursors with a 22 residue pre-peptide. Inclusion of microsomal membranes in translations suggests that the three different pre-peptides each function as co-translationally cleaved signal peptides. However, treatment of processed precursors with endoglycosidase H indicates that co-translational processing of precursors with 22 and 32 residue pre-peptides leads to glycosylation of downstream IGF-I precursor sequences whereas co-translational processing of precursors with 48 residue pre-peptide is not associated with glycosylation.
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PMID:Multiple transcription start sites in the rat insulin-like growth factor-I gene give rise to IGF-I mRNAs that encode different IGF-I precursors and are processed differently in vitro. 827 98

Genomic DNA encoding the 5' region of the porcine IGF-I gene was cloned and sequenced and shown to be highly homologous to that of man, rats and sheep. Two leader exons (exons 1 and 2), which are alternately spliced to exon 3 (encoding part of the mature IGF-I molecule), were identified by RNase protection analysis. In both cases, transcription initiates upstream from exons 1 and 2 at multiple dispersed start sites to yield two distinct IGF-I mRNA transcript classes (1 and 2) which differ in the precursor peptides predicted from their individual leader sequences. The expression of class 1 and 2 transcripts was measured in liver and muscle RNA from two groups of 2-month-old pigs whose energy status had been manipulated within physiological limits to produce marked differences in plasma IGF-I levels and growth rates. For this purpose, RNase protection probes were developed that contained the individual leader exons 1 and 2 linked separately to the common exon 3, so that class-specific and total IGF-I gene expression could be determined in a single assay. At normal plasma IGF-I concentrations (200 ng/ml), class 1 and 2 transcripts comprised 81 and 19% respectively of total liver IGF-I mRNA, while at a lower plasma concentration (90 ng/ml) the corresponding values were 95 and 5% respectively. Although both classes of transcript declined with the decrease in plasma IGF-I, the relative drop in levels of class 2 transcripts (84%) was substantially greater than that of class 1 (54%). In longissimus dorsi, cardiac and soleus muscles IGF-I mRNA was predominantly of class 1 and did not change in response to decreased plasma IGF-I. This suggests that liver-derived endocrine IGF-I has an important function in the regulation of muscle growth and that class 2 IGF-I transcripts are more sensitive to conditions that promote optimal growth.
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PMID:The porcine insulin-like growth factor-I gene: characterization and expression of alternate transcription sites. 829 76

The development of riboprobe expression cassettes for phosphorimager-based quantitation of steady-state transcripts for three different genes using solution hybridization, RNase protection assays is described. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and beta-actin genes are widely used as reporter genes to estimate the amount and integrity of RNA as well as for comparing gene expression among different tissues. To directly compare expression of these two genes in lymphoid tissue and liver, cDNA fragments of beta-actin and GAPDH from both mice and rats were generated by RT-PCR and cloned together into pGEM1 under control of the T7 RNA polymerase promoter. Antisense transcripts from this fusion construct protected the appropriate-sized fragments of beta-actin (115 nt) and GAPDH (214 nt) in RNA isolated from rat spleen, thymus and liver. Expression of GAPDH transcripts was less variable across tissues because this mRNA was only two-fold lower in liver as compared to either thymus or spleen, whereas expression of beta-actin transcripts was eight-fold lower in liver than in these tissues. Two other riboprobe expression cassettes (IGF-I/actin) were constructed by ligating a cDNA fragment of mouse or rat beta-actin that would protect 115 nt to either a mouse or rat IGF-I genomic DNA fragment containing 182 bp of exon 4. These mouse and rat IGF-I/actin riboprobes were used to conclusively demonstrate that rat CSF-1-derived bone marrow macrophages, mouse elicited peritoneal macrophages and the murine PU5-1R macrophage cell line synthesize abundant transcripts for both IGF-I and beta-actin. However, the mouse M1 progenitor myeloid cell line does not express RNA for IGF-I, as demonstrated by the absence of protected transcripts for IGF-I in the presence of abundant protected transcripts for beta-actin. Phosphorimager scanning of the gels revealed that macrophages of both mice and rats express IGF-I transcripts at a level of 60-100% of those found in liver. These data show that a single riboprobe can be developed to generate multigene antisense RNAs that can then be used to quantitatively compare IGF-I transcripts in macrophages and other tissues to an internal standard, with GAPDH transcripts being less variable among tissues than those for beta-actin. This approach should be broadly applicable for measuring a variety of markers of cellular activation.
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PMID:Riboprobe expression cassettes for measuring IGF-I, beta-actin and glyceraldehyde 3-phosphate dehydrogenase transcripts. 830 98

The present studies were designed to investigate the nature of the actions of insulin-like growth factor-II (IGF-II) on granulosa cell steroidogenesis and assess the potential facilitative interactions between IGF-II and other major regulators of ovarian sterol metabolism, viz. estrogen, FSH, and low density lipoprotein (LDL). In serum-free first passage monolayer cultures of swine granulosa cells, human recombinant IGF-II stimulated progesterone production with a half-maximally effective concentration of 4.6 +/- 1.2 ng/ml (0.61 +/- 0.16 nM) between 0-48 h of culture and 27 +/- 5.7 ng/ml (3.6 +/- 0.76 nM) between 48-96 h. Maximal progesterone accumulation increased 12-fold over that in untreated cultures (48-96 h). Over the latter interval, IGF-I stimulated progesterone production approximately 10-fold, with a significantly lower ED50 of 6.1 +/- 0.70 ng/ml (0.78 +/- 0.09 nM; P < 0.01 vs. IGF-II effect). IGF-II (100 ng/ml) enhanced progesterone biosynthesis approximately 2-fold in the presence of 25-hydroxycholesterol, suggesting that IGF-II increases the effective activity of the mitochondrial cholesterol side-chain cleavage enzyme. IGF-II (100 ng/ml) augmented human LDL-promoted progesterone production approximately 18-fold between 0-48 h of culture and approximately 6-fold between 48-96 h. In addition, IGF-II showed time-dependent stimulatory effects on the rates of [125I]iodo-LDL internalization, and the amounts of cell-associated and degraded lipoprotein. IGF-II increased by approximately 10-fold the number of specific high affinity LDL receptors on granulosa cells, with no apparent change in their binding affinity, as assessed in equilibrium competition studies. Coadministration of IGF-II and FSH (100 ng/ml) or estradiol (E2; 1 microgram/ml) for 2 days increased progesterone production synergistically. Cotreatment with FSH or E2 for 4 days decreased the ED50 of IGF-II's stimulation of progesterone accumulation by 61% and 50%, respectively (P < 0.01). Synergistic interactions also existed between IGF-II and 8-bromo-cAMP, which indicates that IGF-II can act in part at cellular loci distal to cAMP generation. Northern blot analysis of total RNA isolated from granulosa cells treated with IGF-II (100 ng/ml), FSH (100 ng/ml), or IGF-II plus FSH for 2 days revealed 5-, 7-, or 8-fold increases, respectively, in the amount of cytochrome P450 cholesterol side-chain cleavage enzyme mRNA. The same treatments produced 6-fold increases in the level of LDL receptor mRNA, as determined by solution hybridization/RNase protection assays.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Mechanisms of regulation of ovarian sterol metabolism by insulin-like growth factor type II: in vitro studies with swine granulosa cells. 834 16

During the menstrual cycle, the endometrium undergoes characteristic changes in response to circulating sex steroids. Intense mitotic activity of glands and stroma occurs in the proliferative (estradiol-dominant) phase, and glandular secretion and stromal differentiation in the secretory (progesterone-dominant) phase. The insulin-like growth factors (IGF-I and IGF-II) promote cellular growth and differentiation and have been proposed to participate in these cyclic endometrial events, acting as mediators of steroid hormones. The objective of this study was to determine whether the messenger RNAs (mRNAs) encoding the IGF peptides and the type I and type II IGF receptors are differentially expressed in human endometrium during the menstrual cycle and in early pregnancy. A solution hybridization ribonuclease protection assay, using 32P-labeled riboprobes for IGF-I, IGF-II, and beta-actin (control), revealed IGF-I gene expression primarily in proliferative and early secretory endometrium and abundant IGF-II gene expression in mid-late secretory endometrium and early pregnancy decidua. Northern analysis, using IGF-I and IGF-II complementary DNA probes, revealed multiple IGF-I mRNAs [2-7.6 kilobase (kb)], expressed primarily in proliferative and early secretory endometrium, and IGF-II mRNAs (1.4-6.0 kb), expressed primarily in secretory endometrium and in early pregnancy decidua. The 7.6-kb IGF-I mRNA and the 6.0-kb IGF-II mRNA were most abundantly expressed. IGF-IEa and IGF-IEb mRNA splicing variants were present in a ratio of about 9:1, respectively. Type I and type II IGF receptor gene expression in endometrium was investigated using specific riboprobes and the ribonuclease protection assay. Messenger RNAs encoding both receptors were more abundantly expressed in the secretory phase and during early pregnancy, compared to the proliferative phase. These results show that mRNAs encoding the IGF peptides and their receptors are differentially expressed in human endometrium, depending on the steroid hormone milieu. The preferential expression of IGF-I mRNA in the proliferative phase supports the hypothesis that IGF-I is an estromedin in human endometrium. The expression of endometrial IGF-II mRNA in the mid to late secretory phase and in early pregnancy supports a role for IGF-II in differentiative functions of the endometrium, perhaps including endometrial tissue shedding in the menstrual cycle or remodeling during early pregnancy.
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PMID:Differential expression of messenger ribonucleic acids encoding insulin-like growth factors and their receptors in human uterine endometrium and decidua. 849

In this study, we examined the expression of insulin-like growth factor (IGF) ligands, receptors, (IGFR1, IGFR2), and binding proteins (IGFBPs) in the human prostate cancer cell line DU145, as well as its mitogenic response to the IGFs. Using RNase protection assays, we found expression of IGF-II, IGFR1, and IGFR2 but failed to detect IGF-I messenger RNA. Distinct binding protein species as well as immunoreactive IGF-II were detected in conditioned media using radioligand and immunoblotting assays. Compared with controls, treatment with exogenous IGF-I and IGF-II resulted in stimulation of monolayer and anchorage-independent growth. Recombinant human IGFBP-1, which binds IGF-II with high affinity, inhibited IGF-II-induced monolayer growth and both baseline and IGF-II-induced anchorage-independent growth in this cell line. Our data suggest IGF-II is as an autocrine growth factor in DU145 cells, and that inhibition of IGF-II-dependent growth of human prostate cancer cells may represent a new therapeutic strategy for this disease.
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PMID:Proliferation of cultured human prostate cancer cells is inhibited by insulin-like growth factor (IGF) binding protein-1: evidence for an IGF-II autocrine growth loop. 853 May 86


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