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)

To study the mechanisms which mediate the transcriptional activation of cardiac genes during alpha adrenergic stimulation, the present study examined the regulated expression of three cardiac genes, a ventricular embryonic gene (atrial natriuretic factor, ANF), a constitutively expressed contractile protein gene (cardiac MLC-2), and a cardiac sodium channel gene. alpha 1-Adrenergic stimulation activates the expression and release of ANF from neonatal ventricular cells. As assessed by RNase protection analyses, treatment with alpha-adrenergic agonists increases the steady-state levels of ANF mRNA by greater than 15-fold. However, a rat cardiac sodium channel gene mRNA is not induced, indicating that alpha-adrenergic stimulation does not lead to an increase in the expression of all cardiac genes. Studies employing a series of rat ANF luciferase and rat MLC-2 luciferase fusion genes identify 315- and 92-base pair cis regulatory sequences within an embryonic gene (ANF) and a constitutively expressed contractile protein gene (MLC-2), respectively, which mediate alpha-adrenergic-inducible gene expression. Transfection of various ANF luciferase reporters into neonatal rat ventricular cells demonstrated that upstream sequences which mediate tissue-specific expression (-3003 to -638) can be segregated from those responsible for inducibility. The lack of inducibility of a cardiac Na+ channel gene, and the segregation of ANF gene sequences which mediate cardiac specific from those which mediate inducible expression, provides further insight into the relationship between muscle-specific and inducible expression during cardiac myocyte hypertrophy. Based on these results, a testable model is proposed for the induction of embryonic cardiac genes and constitutively expressed contractile protein genes and the noninducibility of a subset of cardiac genes during alpha-adrenergic stimulation of neonatal rat ventricular cells.
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PMID:Co-regulation of the atrial natriuretic factor and cardiac myosin light chain-2 genes during alpha-adrenergic stimulation of neonatal rat ventricular cells. Identification of cis sequences within an embryonic and a constitutive contractile protein gene which mediate inducible expression. 185 Apr 19

5'-Flanking sequences from the human atrial natriuretic factor (hANF) gene were subcloned into a reporter plasmid (pSVOCAT) and transfected into primary cultures of neonatal rat atrial cardiocytes. Hybrid hANFCAT genes containing either 2500 or 409 base pairs of 5'-flanking sequence DNA were expressed at similar levels. When sequences between -409 and -332 were deleted, reporter gene (CAT) activity decreased significantly. Expression of the hANFCAT constructs was specific for atrial cells, as no expression was detected in primary cultures of ventricular cardiocytes or nonmyocardial cells derived from the neonatal hearts. Correct transcription start sites for the transfected hANF genes were confirmed by S1 nuclease mapping and RNase protection analysis. A "gel shift" assay was used to identify a specific cardiac nuclear protein which bound to the 5'-flanking sequence of the hANF gene. A 192-base pair PvuII fragment (-400 to -208) associated with a protein in these extracts in a tissue- and sequence-specific fashion. These findings indicate that the DNA sequence between -409 and -332 in the hANF gene harbors a tissue-specific element whose activity may involve association with a cardiac-specific nuclear protein.
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PMID:Upstream sequences confer atrial-specific expression on the human atrial natriuretic factor gene. 296 19

Hammerhead ribozymes targeted to various GUC or GUA sites on rat atrial natriuretic factor (ANF) mRNA were developed. The catalytic activity of ribozymes to four of these sites, synthesized by transcription off synthetic oligodeoxynucleotide duplexes, was studied in detail. In vitro, ribozyme-mediated cleavage was highly Mg(2+)-dependent, and at concentrations approaching those found intracellularly, the rate but not the extent of cleavage was markedly reduced. To test for cellular activity, synthetic genes encoding the ribozymes were cloned between the initiation and termination sequences of the U1snRNA gene or between the T7RNA polymerase promoter and terminator sequences in pSP64. Both constructs had defined initiation and termination sequences to minimize transcript size and for message stability. In vitro the addition of T7 or U1 terminator sequences had variable effects on catalytic activity, presumably due to structural interactions between the ribozyme and the added sequence. The ribozyme-encoding plasmids were cotransfected with an expression plasmid containing a rat ANF cDNA into COS-1 cells using a liposome method, which provided high-level transfection efficiency. Quantitation of ANF mRNA by RNase protection showed marked decreases in ANF transcript levels with both the U1- and the T7-expressed ribozymes directed at three of the four sites on ANF mRNA. With all constructs, target accessibility, determined in vitro, was a more important determinant of intracellular ANF mRNA cleavage than catalytic activity per se. ANF mRNA cleavage was not merely due to an antisense effect, since a mutant construct that was catalytically inactive but could still bind produced less cleavage than the corresponding wild-type ribozyme construct. These findings indicate that both U1 and T7 vector systems provide efficient ribozyme expression for the intracellular cleavage of target mRNA.
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PMID:Functional characterization of ribozymes expressed using U1 and T7 vectors for the intracellular cleavage of ANF mRNA. 791 34

The structure of the mouse natriuretic peptide type-B (BNP) gene was determined by isolating and sequencing genomic clones. The mouse BNP gene was structurally similar to other natriuretic peptide genes and comprised three exons and two introns. Expression of the mouse BNP gene was found only in cardiac tissue as determined by ribonuclease protection analyses. Initiation of transcription was 31 bp downstream from a consensus TATA box as determined by primer extension analysis of cardiac RNA. Comparative DNA sequence analysis identified several DNA elements with potential transcriptional regulatory function. Comparative amino acid sequence analysis showed that the N-terminal portion of the mouse and rat BNP precursors was more conserved than the C-terminal 45-amino-acid sequence that constitute the bioactive BNP-45 peptide. The proteolytic processing site (RXXR-S) generating bioactive BNPs was highly conserved among all BNP precursors and was identical to the consensus site of furin, a calcium-dependent serine endoprotease. Finally, the BNP gene was mapped using recombinant inbred DNA and a polymerase chain reaction-based restriction fragment-length polymorphism assay to mouse chromosome 4 near the atrial natriuretic factor (Anf) locus. No recombination event between Bnp and Anf was evident in the 39 recombinant inbred and inbred strains examined. This physical linkage between the two natriuretic peptide genes expressed in cardiac tissue may be important for their transcriptional regulation.
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PMID:Structure, expression, and genomic mapping of the mouse natriuretic peptide type-B gene. 809 40

The atrial natriuretic peptide (ANP) gene is expressed in several extracardiac tissues where ANP is thought to be involved in autocrine or paracrine regulation. The current studies were designed to characterize the ANP system in rat ovaries. ANP content in rat ovaries was estimated by RIA to be 240 +/- 70 pg/mg protein. HPLC revealed the presence of the 28-amino acid circulating peptide as well as the 126-amino acid prohormone, suggesting that the ovaries are a site of ANP synthesis. Indeed, ANP messenger RNA was detected in this tissue by RNase mapping. ANP present in ovarian extracts displaced [125I]ANP from bovine adrenal receptors (R1 class) in a dose-dependent manner and in parallel to the synthetic peptide, indicating that it possesses biological activity. Immunocytochemical studies localized ANP to interstitial cells surrounding the follicles; weaker but specific staining was also observed in the ovum. High affinity ANP receptors (dissociation constant, 0.30 +/- 0.06 nM; maximum binding capacity, 160 +/- 40 fmol/mg protein) were identified in ovarian membranes. Unlabeled ANP but not c-atrial natriuretic factor (a specific agonist of ANP clearance receptors) competed with binding of [125I]ANP to ovarian membranes in a dose-dependent manner, suggesting that ovarian ANP receptors are predominantly of the R1 class. This was confirmed by cross-linking studies with [125I]ANP, which detected a single protein band with a molecular size of about 120 kilodaltons, corresponding to that of the guanylate cyclase-coupled R1 class of receptor. Consistent with the presence of biologically active receptors, ANP markedly enhanced cGMP accumulation (by 15-fold) in ovarian cells. The presence of both local ANP synthesis and high affinity transducing receptors in the ovaries indicates that the peptide plays a local role in ovarian growth or steroidogenesis.
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PMID:The atrial natriuretic peptide system in rat ovaries. 842 88

The present investigation was designed to (1) determine if atrial natriuretic factor gene expression occurs within invertebrates as well as within vertebrates; (2) determine whether the product of this gene expression is the 126-amino-acid atrial natriuretic factor prohormone or some other molecular species; and (3) evaluate within the same invertebrates if the products of atrial natriuretic factor gene expression are released into their circulation. Utilizing a very sensitive RNase protection assay it was found that atrial natriuretic peptide gene expression occurs within the heart of the oyster, Crassostrea virginica, and within the heart of the blue crab, Callinectes sapidus, but was expressed sevenfold less than in a vertebrate heart (i.e., rat, Rattus norvegiucs). High-performance gel-permeation chromatography followed by N-terminal and C-terminal atrial natriuretic factor prohormone radioimmunoassays indicated that the molecular species synthesized within the oyster and blue crab hearts was the atrial natriuretic factor prohormone. The product(s) of this atrial natriuretic factor gene expression (i.e., atrial natriuretic peptides) was found to be released into the circulation, i.e., hemolymph, of both the oyster and the blue crab.
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PMID:Atrial natriuretic peptide gene expression within invertebrate hearts. 857 60

An antisense oligodeoxynucleotide (ODN) approach was used to investigate whether mitogen-activated protein kinase (MAPK) is necessary for the hypertrophic response in cardiac myocytes. A phosphorothioate-protected 17-mer directed against the initiation of translation sites of the p42 and p44 MAPK isoform mRNAs was introduced into cultured cardiac myocytes by liposomal transfection. At an antisense ODN concentration of 0.2 mumol/L, p42 MAPK protein was reduced by 82% (immunoblot) after 48 hours, and p42 and p44 MAPK activities were reduced by 44% and 60%, respectively. The same concentration of anti-MAPK ODN inhibited development of the morphological features of hypertrophy (sarcomerogenesis, increased cell size) in myocytes exposed to phenylephrine. Phenylephrine-induced activation of the atrial natriuretic factor (ANF) promoter (measured by the activity of a transfected ANF promoter/luciferase reporter gene) and induction of ANF mRNA (measured by RNase protection assay) were also attenuated. We conclude that MAPK is important for the development of the hypertrophic phenotype in this model of hypertrophy.
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PMID:Depletion of mitogen-activated protein kinase using an antisense oligodeoxynucleotide approach downregulates the phenylephrine-induced hypertrophic response in rat cardiac myocytes. 863 45

Genomic sequences encoding mouse C-type natriuretic peptide (CNP) were isolated from bacteriophage libraries and characterized by restriction enzyme and sequence analysis. The mouse CNP gene (Nppc) comprised at least two exons and one intron and included several cis-regulatory elements in the 5'-flanking sequence. The deduced amino acid sequence of mouse CNP-22 was identical to other mammalian CNPs. Analysis of allele distributions in interspecific back-cross and recombinant inbred strains assigned Nppc to chromosome 1. CNP transcripts were detected by ribonuclease protection analysis in brain, ovary, and uterus, with lower levels in testes and epididymus. Uterine CNP transcripts and protein were low in sexually immature mice and adults at estrus and increased at proestrus, but similar variations in ovarian CNP expression were not statistically significant. Atrial natriuretic peptide and B-type natriuretic peptide transcripts were not detected in mouse ovary or uterus. Thus CNP gene expression is regulated by tissue-specific and inducible mechanisms in female reproductive organs. Correlations between CNP expression and uterine fluid content suggest that CNP may regulate uterine fluid balance in mice and other mammals.
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PMID:Isolation, mapping, and regulated expression of the gene encoding mouse C-type natriuretic peptide. 889 53

Following myocardial Infarction (MI) the heart undergoes a process of remodeling characterized by considerable hypertrophy of the non-infarcted myocardium. We have recently characterized the molecular basis of key electrophysiologic alterations that may provide insight into the arrhythmogenecity of post-MI remodeled hypertrophied myocardium. To further characterize other key alterations in the pattern of cardiac gene expression in a time-dependent manner, we have measured mRNA and immunoreactive protein levels of selective cardiac genes in the remodeled hypertrophied left-ventricular (LV) myocardium of rats, 3 and 21 days after left-coronary ligation and compared the results with sham-operated rats. RNase protection assay was performed to assess the expression of c-fos, atrial natriuretic factor (ANF), brain natriuretic factor (BNF), alpha2/3 isoform of Na-K ATPase, cardiac alpha/beta isoform of myosin heavy chain (MHC). Compared to the sham group, the expression of c-fos was increased 10-fold (P<0.02) in the MI group on day 3, but unlike other overload hypertrophy models, the expression remained elevated by three-fold on day 21. Similar to other overload models, the ANF and BNF expression increased significantly. No alterations were observed in the expression of cardiac alpha-actin. There was reexpression of the fetal isogene form of MHC and Na-K ATPase after MI. The beta-MHC mRNA levels, the fetal isoform of MHC, returned to basal levels after 21 days. After an initial five-fold decrease the adult isoform of alphaNa-K ATPase, alpha2 Na-K ATPase mRNA, returned to control levels and similar changes were seen in the corresponding protein levels. These findings indicate that during LV remodeling and hypertrophy following MI, there is an upregulation of early response genes and fetal isogene expression. The pattern of activation, however, is distinct from that observed in other overload models, indicating the possible involvement of alternate signal transduction pathways.
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PMID:Alterations in cardiac gene expression during ventricular remodeling following experimental myocardial infarction. 951 38

Left ventricular hypertrophy (LVH) entails numerous functional and molecular changes that ultimately lead to cardiac insufficiency. The renin-angiotensin system and adrenergic receptor signalling pathway have both been implicated in LVH progression and interactions between these factors may precipitate contractile dysfunction. We therefore investigated cardiac function in hypertensive rats transgenic for the human renin and angiotensinogen genes (TGR) having a genetic activation of the renin-angiotensin system, stroke-prone spontaneously hypertensive rats (SHR) and normotensive controls (CTR) aged 6 weeks. The isolated perfused heart model was used and the effect of isoproterenol (0.1-1000 nmol/L on cardiac function was studied. Cardiac protein and gene expression was studied by Western blot and RNase protection assay. TGR had 75 mmHg higher blood pressure and a 24% higher cardiac/body weight ratio than CTR; blood pressure in SHR was 17 mmHg higher without heart weight difference (p < 0.05). Basal Pmax, +dP/dt and -dP/dt were higher in TGR and SHR compared with CTR hearts. Isoproterenol stimulated these parameters by a maximum factor 6-8 in CTR and SHR but had almost no effect in TGR (p < 0.05). Basal CF per g heart weight was similar in all experimental groups. Isoproterenol produced a significantly smaller vasodilation in TGR compared with CTR or SHR. beta 1 and beta 2 receptor and Gs alpha proteins were similar in TGR, SHR and CTR. Gi alpha was increased in TGR hearts (p < 0.05). Converting enzyme and atrial natriuretic factor mRNA expression was increased (p < 0.01) while beta 1 receptor, adenylyl-cyclase V, SERCA2a and phospholamban mRNA expression was unchanged in TGR compared with CTR. Thus, LVH in TGR is characterised by early adrenergic dysfunction and beta 1 receptor signalling abnormalities indicating progressive functional deterioration. The data may serve as support for an early preventive intervention in angiotensin-II dependent cardiac hypertrophy and may have also implications for patients with genetic alterations of the renin-angiotensin system.
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PMID:[A comparative study of cardiac function in transgenic hypertensive rats, in spontaneously hypertensive rats and in normotensive rats]. 1098 44


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