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)

Fibronectin expression was shown recently to increase in the rat aorta in response to experimental hypertension. Fibronectin is known to alter the phenotype of vascular smooth muscle and endothelial cells, and relative changes in the expression of different isoforms of fibronectin, generated by alternative splicing and distinguished by the absence or presence of inserts designated as EIIIA, EIIIB, and V, may reflect a change in cell phenotype. In the present study we examined the expression of alternatively spliced forms of aortic fibronectin during deoxycorticosterone-salt hypertension. Aortic RNA was analyzed quantitatively using Northern blot analysis and ribonuclease protection assays. Using Northern blot analysis, deoxycorticosterone-salt treatment for 21 days led to a 4.9-fold increase in EIIIA fibronectin messenger RNA, while EIIIB and V forms increased by 2.6- and 2.5-fold, respectively. As determined by ribonuclease protection assays, the percentage of fibronectin transcripts containing either EIIIA, EIIIB, or V in control aorta was 7.3%, 19%, and 40%, respectively. The percentage of EIIIA transcripts increased 42% over control levels after 21 days of deoxycorticosterone-salt treatment, whereas no proportionate change in the other alternatively spliced forms was found. Thus, all forms increased, but a selective increase in the EIIIA form was induced. Analogous increases in each of the fibronectin isoforms were found in the spontaneously hypertensive rats when compared with age-matched Wistar-Kyoto or Wistar rats, and 40-week-old animals showed increases over 10-week-old animals in all strains, consistent with an age-dependent increase in aortic fibronectin expression.
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PMID:Hypertension induces alternatively spliced forms of fibronectin in rat aorta. 161 48

Advanced glycosylation endproducts (AGEs) are derived from the nonenzymatic addition of glucose to proteins. AGEs have been found to accumulate on tissue proteins in patients with diabetes, and their accumulation is thought to play a role in the development of diabetic complications. The finding that macrophages and endothelial cells contain AGE-specific receptors led us to examine whether mesangial cells (MCs) also possess a mechanism for recognizing and processing AGEs. Membrane extracts isolated from rat and human MCs were found to bind AGE-bovine serum albumin (BSA) in a saturable fashion, with a binding affinity of 2.0 +/- 0.4 x 10(6) M-1 (500 nM). The binding was specific for the AGE adduct, since AGE-modified collagen I and ribonuclease both competitively inhibited 125I-AGE-BSA binding to MC membranes, while the unmodified proteins did not compete. Binding of AGE proteins was followed by slow internalization and degradation of the ligand. Ligand blotting of MC membrane extracts demonstrated three distinct AGE-binding membrane proteins of 50, 40, and 30 kD. Growth of MCs on various AGE-modified matrix proteins resulted in alterations in MC function, as demonstrated by enhanced production of fibronectin and decreased proliferation. These results point to the potential role that the interaction of AGE-modified proteins with MCs may play in vivo in promoting diabetic kidney disease.
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PMID:Human and rat mesangial cell receptors for glucose-modified proteins: potential role in kidney tissue remodelling and diabetic nephropathy. 165 49

We studied expression of laminin, fibronectin, and Type IV collagen in the testis by means of immunofluorescence and immunoblot analysis and also examined gene expression of fibronectin using the ribonuclease protection assay. By immunofluorescence on sections from 20-day-old rats, laminin, fibronectin, and Type IV collagen were found in the basement membrane of the seminiferous tubules and in the interstitial regions of the testis. No localization of any extracellular matrix components was found inside the sectioned cells. However, when Sertoli cells were cultured on glass coverslips, laminin and Type IV collagen were both found inside the cells, suggesting new synthesis. In cultured peritubular cells, Type IV collagen, laminin, and fibronectin were found within the cells. When examined by immunoblot analysis, freshly isolated Sertoli and peritubular cells from 20-day-old rats did not demonstrate production of laminin or fibronectin. After 5 days in culture, peritubular cells produced both laminin and fibronectin, whereas cultured Sertoli cells produced only laminin. In contrast, freshly isolated and cultured Sertoli and peritubular cells all produced Type IV collagen. Moreover, the ribonuclease protection assay indicated that the bulk of fibronectin gene expression occurs within the first 10 days of postnatal development, with lower maintenance levels occurring thereafter. These results indicate that in the testis the highest levels of expression of laminin and fibronectin occur during development and in primary cell culture, whereas expression of Type IV collagen is higher at later stages.
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PMID:Differential expression of extracellular matrix components in rat Sertoli cells. 170 45

Fibronectin isoforms are generated by the alternative splicing of a primary transcript derived from a single gene. In rat at least three regions of the molecule are involved: EIIIA, EIIIB, and V. This study investigated the splicing patterns of these regions during development and aging, by means of ribonuclease protection analysis. Between fetal and adult rat, the extent of inclusion of the EIIIA and/or EIIIB region in fibronectin mRNA varied according to the type of tissue analyzed; but the inclusion of the V region, and in particular the V25 alternative variant, was significantly higher in all fetal than in adult tissues. These data suggest a crucial role of the V25 variant, possibly related to its interaction with the alpha 4 beta 1 integrin receptor during development. On the other hand, during aging, the only significant change observed in the splicing pattern was a decrease in the EIIIA variant in brain. The high inclusion levels of the EIIIA and EIIIB regions in young adult brain suggest that these segments may play an important role in differentiated brain tissue. The decreasing levels of inclusion of the EIIIA segment in brain fibronectin mRNA during aging may be an age-related marker with functional consequences.
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PMID:Tissue-specific splicing pattern of fibronectin messenger RNA precursor during development and aging in rat. 204 Jun 49

The primary gene transcript for the adhesive extracellular matrix glycoprotein fibronectin (FN) is alternatively spliced in three regions (EIIIA, EIIIB and V). At least one of these regions (V) has been shown to encode cell-binding sites, suggesting that splicing represents a mechanism to create functionally different forms of FN at different times and places. In order to test this hypothesis, we have examined the extent of alternative splicing of fibronectin during embryonic development. The distribution of the different spliced forms of FN mRNA in developing chicken embryos was determined using probes specific for the spliced regions in ribonuclease protection and in situ hybridization experiments. At embryonic day 2-4 (E2-4), all three spliced regions were included wherever FN mRNA was detected. At E16, however, we found spatially distinct splicing differences within the embryo, with cell-type-specific splicing excluding EIIIA and/or EIIIB in some tissues. In contrast, we did not detect exclusion of the V region. In a more detailed developmental study of the simplest of these tissues, the chorioallantoic membrane, we found that EIIIB was preferentially excluded after the completion of growth. These results suggest that FN splicing is used during development as a mechanism to create different forms of FN within the extracellular matrix by the inclusion or exclusion of specific segments. The data are consistent with an essential role for one of these segments, EIIIB, in the migration and/or proliferation of embryonic cells prior to their terminal differentiation and also suggest possible roles for the EIIIA segment.
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PMID:Alternative splicing of fibronectin is temporally and spatially regulated in the chicken embryo. 259 21

Insulin-like growth factor binding proteins (IGFBPs) constitute a family of at least six secreted proteins that bind insulin-like growth factors I and II (IGF-I and -II) and are capable of modifying IGF actions on target cells. We previously have purified an approximately 29-kDa IGFBP that is secreted by myoblasts during their terminal differentiation, have identified the protein as mouse IGFBP-5, and have cloned its cDNA (James et al., 1993). In this study, we have characterized the mouse IGFBP-5 gene, established its pattern of expression in the adult mouse, and defined its chromosomal location. The 17-kb gene was isolated on overlapping cosmid and lambda clones, and the four exons encoding the 5914-bp mRNA were sequenced. The 5' end of the gene was mapped by solution-hybridization ribonuclease protection assay to two discrete sites in exon 1 that were separated by 21 bp. The relative use of each transcription start site was found to vary among different mouse tissues. By interspecies backcross mapping using progeny derived from matings of [(C57BL/6J X Mus spretus)F1 x C57BL/6J] mice, the IGFBP-5 gene was localized to the proximal region of chromosome 1 in tight linkage with fibronectin 1 (0 recombinants in 168 mice analyzed). Since this part of chromosome 1 shares homology with human chromosome 2q, and since fibronectin has been mapped to 2q34-q36, it is likely that human IGFBP-5 will reside on 2q as well. Characterization of the mouse IGFBP-5 gene now provides a starting point for studying the roles and regulation of this protein in development.
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PMID:Organization, expression, and chromosomal location of the mouse insulin-like growth factor binding protein 5 gene. 751 10

Fibronectin mRNAs that include the alternatively spliced exons EIIIA, EIIIB, and V are prevalent during embryogenesis, and EIIIA and EIIIB reappear during wound healing. Using ribonuclease protection analyses, we found an up-regulation of V120 (containing the alpha 4 beta 1 integrin binding site), as well as EIIIA, and EIIIB in fibronectin mRNAs in the crush-injured adult rat sciatic nerve. In situ hybridization using splice variant-specific probes revealed that cells within endoneurial tubes of the injured nerve synthesize these embryonic forms of fibronectin. Our results suggest that embryonic fibronectins synthesized within the nerve contribute to the permissiveness of the peripheral nervous system to axon regrowth and a mechanism by which alternative splicing of the V region in fibronectin mRNA could enhance nervous system regeneration.
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PMID:Embryonic fibronectins are up-regulated following peripheral nerve injury in rats. 770 41

The temporal changes in the expression of fibronectin and other extracellular matrix genes were studied in rat aortic rings incubated in vitro in a serum-free medium. Changes in all forms of fibronectin mRNA increased progressively during the 24-hour incubation period, although an increase in the alternatively spliced form of fibronectin designated EIIIA was most pronounced. Both collagen and elastin mRNA levels decreased markedly during the 24-hour interval, as did alpha-actin mRNA. The increase in the relative amount of the EIIIA isoform after a 24-hour incubation was also shown using ribonuclease protection assays. In situ hybridization showed the distribution of the induced fibronectin mRNA to be within all cell types, including endothelial cells, medial smooth muscle cells, and adventitial fibroblasts. Localization in the media was not uniform and was clearly identified mainly in clusters of cells distributed throughout the media. The early induction of fibronectin mRNA was inhibited by genistein, implicating tyrosine kinase activation as a causative factor in fibronectin expression. The in vitro changes reported may reflect a phenotypic change in vascular cell types that is both similar to and different from the changes reported in vivo under conditions in which vascular injury and repair occur.
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PMID:Selective induction of an embryonic fibronectin isoform in the rat aorta in vitro. 837 Jan 23

Different mRNAs for fibronectin arise from the variable processing of a single primary transcript. We used ribonuclease protection assay to investigate the changes occurring in fibronectin expression and the alternative splicing of mRNA precursor during aging in vitro of human diploid endothelial cells. Senescent endothelial cells release more protein and contain 4-5-fold more fibronectin mRNA than young cells. The pattern of alternative splicing of fibronectin mRNA, with the EDA and the CS1 segments largely included (35% and 77%, respectively) and the EDB segment undetectable, correlates well with previous studies at the protein level both in vitro and in vivo. No changes in the splicing pattern of fibronectin mRNA precursor were detected during endothelial cellular senescence. The increased expression of fibronectin in senescent cells may be a result of the activity of interleukin-1 alpha, which is overexpressed in senescent endothelial cells. It could be also important in vivo during aging and in atherosclerotic lesions.
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PMID:Expression and alternative splicing of fibronectin mRNA in human diploid endothelial cells during aging in vitro. 850 66

The Milli-Q PF Plus water polishing system is equipped with high-purity ion and organic removal media and a capillary fiber ultrafiltration device. The system produces ultrapure water practically free of ribonuclease contamination. The necessity for diethyl pyrocarbonate (DEPC) treated solutions in RNA molecular biological procedures was tested by preparing RNA from a variety of tissues and tissue cultured cells using either DEPC-treated, autoclaved solutions or pure Milli-Q PF water dispensed directly from the system. Tissue sources included rabbit brain, heart, lung, liver, kidney, and bladder as well as cultured human corpus cavernosum smooth muscle cells (HCCSMC). RNA was prepared by solubilization in guanidinium isothiocyanate, phenol/chloroform extraction, and isopropanol precipitation followed by Northern blot analysis. Hybridization with fibronectin (approximately 7.6kb) and glyceraldehyde-3-phosphate dehydrogenase (1.2kb) revealed that water from a Milli-Q PF water system performed as well as DEPC-treated, autoclaved solutions. RNA stability at 37 degrees C was examined for various times using rabbit lung RNA in either DEPC-treated water or Milli-Q PF water. Intact RNA was detected after 6 hours in total RNA and by Northern blots hybridized with fibronectin. There was no significant difference in RNA degradation between DEPC-treated water or Milli-Q PF water. We conclude that Milli-Q PF water is an acceptable substitute to DEPC-treated water for the preparation of RNA and Northern blot analysis.
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PMID:Comparison of Milli-Q PF plus water with DEPC-treated water in the preparation and analysis of RNA. 864 47


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