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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The relative steady-state abundance of cardiac and skeletal alpha-actin mRNAs at different stages of embryonic skeletal and cardiac (striated) muscle development was determined by a reverse transcriptase extension assay employing an single oligonucleotide primer complementary to a perfectly conserved region near the 5' end of both mRNAs. Both mRNAs were found to be present at every stage of embryonic striated muscle development tested, including the earliest assayable stages of limb muscle and cardiac muscle development. At early stages of skeletal muscle development the two mRNAs are present at similar levels while at later stages the abundance of the skeletal alpha-actin mRNA far exceeds that of the cardiac alpha-actin mRNA. Both mRNAs are also present at similar levels throughout embryonic cardiac muscle development while in adult cardiac muscle the cardiac alpha-actin mRNA predominates over the skeletal alpha-actin mRNA. These results for early embryonic striated muscle, in combination with previous results with late embryonic and adult striated muscle, indicate that both genes are coexpressed throughout striated muscle ontogeny. These two genes may not, therefore, be regulated under unique tissue-specific regulatory programs but each may have acquired regulatory elements which confer important quantitative differences in their level of expression in mature striated muscle cells.
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PMID:The skeletal and cardiac alpha-actin genes are coexpressed in early embryonic striated muscle. 375 81

Visna-maedi virus induces in sheep an interstitial lung disease characterised by an accumulation of smooth muscle cells (SMC) or myomatosis. Infection by HIV-1 has been recently associated with disorders of the vessel-derived cells: primary pulmonary hypertension, coronary artery disease and smooth muscle tumors in humans. We hypothesized that, besides their regular targets (i.e. macrophages and lymphocytes), lentiviruses could infect smooth muscle cells. Smooth muscle cell cultures derived from ovine aorta were infected with visna-maedi virus strain K1514. The cultured cells were smooth muscle cells as demonstrated by their antigenic expression of alpha-actin and vimentin. The lentiviral infection of the smooth muscle cells was demonstrated by a typical cytopathic effect (syncytia), the expression of virus specific antigens, and the presence of genomic RNA detected by Northern blot analysis and RT PCR. The detection of a reverse transcriptase activity, the presence of viral RNA in supernatants of infected smooth muscle cells detected by RT PCR and their ability to infect ovine permissive fibroblasts demonstrated a productive infection. The ability of smooth muscle cells to be infected by lentiviruses may participate in the pathogenesis of the tissue damage associated with the lentiviruses such as myomatosis in sheep and vascular disease in humans.
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PMID:Ovine aortic smooth muscle cells allow the replication of visna-maedi virus in vitro. 754 8

In the present study, we have used single chicken blastoderms of defined early developmental stages, beginning with the prestreak stage, stage 1 (V. Hamburger and H. L. Hamilton, J. Morphol. 88:49-92, 1951), to analyze the onset of cardiac myogenesis by monitoring the appearance of selected cardiac muscle tissue-specific gene transcripts and the functional expression of the myocyte enhancer factor 2 (MEF-2) proteins. Using gene-specific oligonucleotide primers in reverse transcriptase PCR assay, we have demonstrated that the cardiac myosin light-chain 2 (MLC2) and alpha-actin gene transcripts appear as early as stage 5, i.e., immediately after the cardiogenic fate assignment at stage 4. Consistent with this observation is the developmental expression pattern of DNA-binding activity of BBF-1, a cardiac muscle-specific member of the MEF-2 protein family, which also begins at stage 5 prior to MEF-2. Differential expression of DNA-binding complexes is also observed with another AT-rich DNA sequence (CArG box) as probe, but the binding pattern with the ubiquitous TATA-binding proteins remains unchanged during the same developmental period. Thus, the cardiogenic commitment and differentiation of the precardiac mesoderm, as exemplified by the appearance of cardiac MEF-2, MLC2, and alpha-actin gene products, occur earlier than previously thought and appear to be closely linked. The onset of skeletal myogenic program follows that of the cardiogenic program with the appearance of skeletal MLC2 at stage 8. We also observed that mRNA for the MEF-2 family of proteins appears as early as stage 2 and that for CMD-1, the chicken counterpart of MyoD, appears at stage 5. The temporal separation of activation of cardiac and skeletal MLC2 genes, which appears immediately after the respective fate assignments, and those of cardiac MEF-2 and CMD-1, which occur before, are consistent with the established appearance of the myogenic programs and with the acquisition pattern of the two tissue-specific morphological characteristics in the early embryo. The preferential appearance of BBF-1 activity in precardiac moesderm, relative to that of MEF-2, indicates that these two protein factors are distinct members of the MEF-2 family and provides a compelling argument in support of the potential role of BBF-1 as a regulator of the cardiogenic cell lineage determination, while cardiac MEF-2 might be involved in maintenance of the cardiac differentiative state.
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PMID:Differential expression of the myocyte enhancer factor 2 family of transcription factors in development: the cardiac factor BBF-1 is an early marker for cardiogenesis. 803 95

PTH-related peptide (PTHrP), the factor mediating the syndrome of humoral hypercalcemia of malignancy, is also expressed in smooth muscle cells (SMC) of the urinary bladder and uterus in response to mechanical distention and fetal occupancy, respectively. Vascular SMC also produce PTHrP, and its expression is induced by serum and vasoconstrictors, such as angiotensin-II. To determine whether mechanical distension affected vascular PTHrP gene expression, the abdominal aorta of adult male rats was balloon-distended, and aortae were collected at various times after the intervention. PTHrP mRNA was determined by competitive reverse transcriptase-polymerase chain reaction, using sequential dilutions of a cloned internally truncated PTHrP RNA fragment as standard. The molar concentration of PTHrP mRNA was obtained by extrapolating at a standard/wild-type band intensity ratio of 1:1. Aortic PTHrP mRNA was induced from a basal level of 19, to 22, 46, 36, 13, 12, 22, and 20 attamoles/mg total RNA 1, 2, 12, 24, and 48 h and 7 and 48 days after balloon distension, respectively. To determine whether mechanical events directly regulate vascular PTHrP gene expression, primary rat aortic SMC were plated and placed on a rocking device at 20 oscillations/min to create a gentle flowing motion of the culture medium. Rocking induced PTHrP mRNA of SMC exposed to either serum-free medium or 10% serum by 2.5-and 4.0-fold at 4 h, and 2.9- and 3.7-fold at 24 h, respectively. These effects were oscillation rate dependent, potentiated by angiotensin-II, and specific, as similar changes were not observed in alpha-actin mRNA content. Flow motion-induced PTHrP mRNA at 24 h was partially decreased by 10(-6) M colchicine (which inhibits microtubule assembly), but not by cytochalasin-E (which disrupts actin polymerization). As PTHrP is a known vasorelaxant, we propose that mechanical events induce the release of PTHrP by SMC, possibly to serve as a compliance factor or an agent for vascular remodeling.
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PMID:Mechanical stimuli induce vascular parathyroid hormone-related protein gene expression in vivo and in vitro. 815 26

We examined the transcriptional activity profile of the gene for atrial natriuretic factor (ANF) in mouse embryonal carcinoma P19 cells which had been induced for in vitro cardiac myogenesis. Differentiation was assessed visually, by the degree of spontaneous beating activity, and by the appearance of striated muscle structures detected by immunofluorescence with a myosin heavy chain antibody. Northern blot analysis of RNA isolated at regular intervals throughout the differentiation program revealed abundant cardiac alpha-actin transcripts beginning at Day 6, reaching maximum levels during Days 7 to 8 and declining to low levels by Days 12 to 15. Throughout this period, the transcriptional profile of the ANF gene was similar to that of alpha-actin but at lower levels; thus, in vivo stages of abundant ANF and structural muscle gene transcription were not reached and these gene expression states appear to be uncoupled. Using the more sensitive assay of reverse transcriptase-mediated polymerase chain reactions, we observed the presence of ANF transcripts even in small samples of muscle-induced P19 cells and not in neuron-induced or undifferentiated P19 cells. Induced ANF transcript levels reached about 5-10% that found in adult atrium muscle tissue. ANF gene activity was further corroborated by nuclear transcriptional run-on assays. The P19 stem cell model system will be of value in the study of early events during cardiac muscle commitment and differentiation.
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PMID:Activation of the gene for atrial natriuretic factor during in vitro cardiac myogenesis by P19 embryonal carcinoma cells. 834 90

Secretory nonpancreatic type II phospholipase A2 (snpPLA2) hydrolyzes fatty acids at the sn-2 position in phospholipids releasing free fatty acids (FFAs) and lysophospholipids. These products may act as intracellular second messengers or can be further metabolized into proinflammatory lipid mediators. The presence of snpPLA2 in extracellular fluids and serum during inflammation has suggested a role of the enzyme in this process. However, the presence of snpPLA2 in a variety of normal tissues suggests that snpPLA2 may also have physiological functions. Atherosclerosis appears to have an inflammatory component. Here we report on the snpPLA2 localization in normal and atherosclerotic lesions and on the properties of the isolated enzyme. A strong snpPLA2 immunoreactivity was observed in the arterial media that was colocalized with alpha-actin-positive vascular smooth muscle cells (SMCs) in both normal and atherosclerotic vessels. In aortic atherosclerotic lesions, snpPLA2 was observed colocalized with CD68-positive macrophages and HHF-35-positive SMCs and extracellularly in the lipid core. snpPLA2 was isolated from human normal arteries and from aorta with lesions. The enzyme was isolated by acid extraction of normal arterial tissues followed by immunoaffinity chromatography. The purified snpPLA2 had an expected molecular weight of 14 kD by polyacrylamide gel electrophoresis and appeared as a single band in immunoblotting. The enzymatic activity was followed by measuring release of fatty acids from phospholipid liposomes or LDL as substrates. The enzymatic activity was inhibited with two specific inhibitors for human snpPLA2: (1) monoclonal antibody 187 and (2) LY311727, a synthetic selective inhibitor. The mRNA for snpPLA2 was detected with reverse transcriptase polymerase chain reaction. These results indicate that snpPLA2 is present in human arteries and that it is able to hydrolyze phospholipids in LDL. The results support the hypothesis that snpPLA2 can release proinflammatory lipids at places of LDL deposition in the arterial wall.
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PMID:Localization of nonpancreatic secretory phospholipase A2 in normal and atherosclerotic arteries. Activity of the isolated enzyme on low-density lipoproteins. 908 85

1. Fast-twitch skeletal muscle fibres were enzymatically dissociated from adult mouse flexor digitorum brevis (FDB) muscles and maintained in culture without or with chronic low frequency stimulation (one 5 s train of 5 Hz pulses per minute) for up to 6 days. Single fibre reverse transcriptase-polymerase chain reaction was conducted to coamplify beta-myosin heavy chain (beta-MHC) and alpha-skeletal actin mRNA from the same fibre. 2. Chronic low frequency electrical stimulation of FDB fibres in culture increased the level of mRNA for beta-MHC. In unstimulated fibres there was a slight decline in the beta-MHC mRNA level. As an internal control there was no increase in the level of mRNA for alpha-actin in the identical individual stimulated or unstimulated fibres. 3. Neither the percentage of fibres exhibiting beta-MHC protein nor the Ca2+ transients recorded from individual fibres subjected to the same pattern of stimulation showed any difference between stimulated and unstimulated fibres over the period in culture. 4. This system provides a convenient in vitro model system for studying activity-dependent control of fibre type-specific gene expression in adult skeletal muscle fibres in culture.
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PMID:Fibre type-specific gene expression activated by chronic electrical stimulation of adult mouse skeletal muscle fibres in culture. 976 24

Skeletal or smooth muscle alpha-actin isoform is reexpressed in hypertrophied or failing heart in experimental animals, however, it has been unknown whether there is a switching of these genes in the human heart. In this study, we assessed alpha-actin isoform mRNA in biopsied specimens from the left ventricle of patients with dilated cardiomyopathy (DCM). The biopsy was performed in 21 patients (DCM; 16, normal; 5). Skeletal and smooth muscle alpha-actin mRNA levels were assessed by quantitative reverse transcriptase-polymerase chain reaction using a synthetic RNA as internal control. The level of both alpha-actin mRNAs correlated well with the mean cardiomyocyte diameter as an index of cardiac hypertrophy. Further, there was a clear correlation between the expression of skeletal alpha-actin mRNA or smooth muscle alpha-actin mRNA and the ejection fraction. These results suggested that mRNAs for skeletal and smooth muscle alpha-actin isoforms are increased in the hypertrophic and/or failing heart. These mRNAs may be an useful marker for the pathological state of cardiomyopathy.
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PMID:Skeletal and smooth muscle alpha-actin mRNA in endomyocardial biopsy samples of dilated cardiomyopathy patients. 982 Jan 22

Previous studies of neointima formation on Dacron vascular grafts mainly focused on the late stages using immunohistochemistry staining for von Willebrand factor (vWF) and smooth muscle (SM) alpha-actin. However, it is impossible to use immunohistochemistry to study the early events of neointima formation, because graft samples lack sufficient cellular material. Therefore, we used reverse transcriptase-polymerase chain reaction (RT-PCR) to demonstrate dynamic changes of SM and endothelial markers during the early stages of neointima formation. Preclotted Dacron grafts were implanted in the descending thoracic aorta of 14 mongrel dogs. Specimens were retrieved at 1-4 weeks. Total RNAs were extracted from mid-portion of graft flow surfaces, and RT-PCR for vWF, SM myosin heavy chain (MHC), and SM alpha-actin were performed and expressed as a ratio to the ribosome s17 signal. SM MHC and vWF mRNA expression was low at 1-2 weeks but elevated at 3-4 weeks (P < 0.05). However, SM alpha-actin mRNA levels were expressed consistently throughout the study period. At 3-4 weeks, vWF mRNA expression was inversely correlated to thrombus formation on the graft flow surface. Increased expressions of SM MHC and vWF mRNA corresponded to the formation of neointima and an endothelial layer at the later stages. However, SM alpha-actin mRNA expression did not vary during the healing process. The application of RT-PCR should permit further studies of gene regulation in the early vascular graft healing process in vivo. This model can also be used to study the molecular events that are involved in SM cell differentiation.
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PMID:Dynamic Changes of Smooth Muscle and Endothelial Markers in the Early Healing Process of Dacron Vascular Grafts in the Dog, Using RT-PCR. 1075 7

We previously described the cloning of the late gestation lung 1 gene (LGL1), a novel glucocorticoid-inducible gene expressed in the mesenchyme of fetal lung. We report here evidence for a role of the LGL1 gene product (lgl1) in fetal rat lung airway branching morphogenesis, temporal and spatial localization of LGL1 mRNA and lgl1 protein in fetal rat lung, and a correction of the previously published LGL1 sequence. Both the mRNA and protein were detected during fetal lung development. LGL1 mRNA was detected from gestational Day 12 by reverse transcriptase-polymerase chain reaction, and from Day 13 by in situ hybridization. lgl1 protein was detected from Day 18 by Western analysis and from Day 16 by immunohistochemistry. The types of cells expressing LGL1 mRNA and lgl1 protein were assessed by immunohistochemical staining of adjacent serial tissue sections for markers of mesenchymal (vimentin) and smooth muscle (alpha-actin) cells. As gestation advanced, increasing amounts of mRNA and protein were expressed in these cells. In support of a role for lgl1 in airway branching morphogenesis, antisense (but neither sense nor scrambled) oligodeoxynucleotides directed against LGL1 inhibited airway branching in fetal rat lung buds in explant culture, in a dose- and time-dependent manner. The levels of lgl1 protein and LGL1 mRNA expression were decreased in those explants that had inhibited airway branching, compared with the uninhibited controls. Our findings suggest that lgl1 plays an important role in fetal airway branching morphogenesis.
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PMID:Antisense oligodeoxynucleotides decrease LGL1 mRNA and protein levels and inhibit branching morphogenesis in fetal rat lung. 1254 Apr 91


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