Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ability of TSH to stimulate synthesis and release of thyroid hormones in ovine thyroid follicles in vitro depends partially on a synergy with insulin-like growth factors (IGFs). The cellular availability of IGFs may be influenced by the release of several IGF binding proteins (IGFBPs). The purposes of these studies was to 1) further characterize the species of IGFBPs synthesized by thyroid follicles, 2) examine the ability of TSH and cortisol to alter IGFBP gene expression and protein release, and 3) investigate the actions of exogenous IGFBPs on thyroid cell function. Adult sheep thyroid follicles were isolated after collagenase digestion, grown to confluence in Coon's modified Ham's F12M medium (OH) with the addition of transferrin, glycylhistidyl-lysine, somatostatin (3H), cortisol and insulin, and maintained in serum-free test media with or without further supplements for up to 48 h. Conditioned media were analyzed for IGFBP presence by Western ligand blotting, and by immunoblotting using specific antisera against bovine IGFBP-2 and human IGFBP-5. IGFBP mRNAs from follicles were identified by Northern blot hybridization using [32P]labeled complementary DNAs encoding ovine IGFBP-1 or -2, and rat IGFBP-4, -5, or -6. Uptake and organification of Na[125I] were measured by incorporation into trichloroacetic acid-precipitable material. Isolated thyroid follicles synthesized four species of IGFBPs in either 0H or 3H medium as detected by ligand blotting, of sizes 40-46, 34, 28, and 18 kilodaltons (kDa), respectively. The 32 kDa IGFBP was identified immunologically as IGFBP-2, whereas the 28 kDa and 18 kDa species were identified as IGFBP-5. Northern blot hybridization of total RNA from cells in 3H medium demonstrated an IGFBP-2 messenger RNA (mRNA) [1.4 kilobase (kb)], an IGFBP-4 mRNA (2.6 kb), and two IGFBP-5 mRNAs (6 kb and 1.8 kb). No IGFBP-1 or -6 mRNAs were detected. Incubation of cultured follicles with TSH (30-500 microU/ml) caused a dose-dependent decrease in the abundance of all IGFBP mRNAs and released proteins, which were reduced further by TSH together with cortisol (10 nM). When the inhibitory effect of TSH and cortisol was removed, IGFBP-2 mRNA increased within 3 h and was 7-fold greater within 12 h. IGFBP-2 did not appear in the conditioned medium until 12 h after TSH removal, along with the other IGFBP species.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Hormonal regulation and biological actions of insulin-like growth factor binding proteins in isolated ovine thyroid follicles. 750 36

Insulin-like growth factor (IGF)-binding protein-5 (IGFBP-5) is uniquely regulated throughout MC3T3-E1 osteoblast differentiation: IGFBP-5 is first detectable in conditioned medium (CM) of replicating preosteoblasts (day 5); IGFBP-5 levels peak between culture days 8-12, then decline to almost undetectable levels in mature osteoblast cultures (> day 18) despite the persistence of IGFBP-5 messenger RNA. These observations suggest that IGFBP-5 concentrations may be regulated by posttranslational mechanisms. To determine whether proteolysis contributes to the disappearance of IGFBP-5 in CM of mature osteoblasts, serial samples of MC3T3-E1 cell CM obtained during a 30-day culture period were analyzed for IGFBP-5-degrading protease activity. Using [125I]recombinant human IGFBP-5 substrate zymography, we demonstrated that proteases with M(r) of 52-72 and 97 kilodaltons (kDa) were present in CM, and protease activity increased in concentration as cultures matured. The 52- to 72-kDa proteases were cation dependent and were inhibited by tissue inhibitor of metalloproteinase 1, a specific inhibitor of matrix metalloproteinases (MMPs), identifying them as MMPs. Furthermore, antisera to human MMP-1 and -2 immunoprecipitated IGFBP-5-degrading proteases with M(r) of 52 and 69/72 kDa, respectively, suggesting that homologous murine MMPs degrade IGFBP-5. Finally, MC3T3-E1 cell CM contained immunoreactive MMP-1 and -2, and MMP-2, in particular, increased significantly throughout differentiation. In contrast, the 97-kDa protease was neither inhibited by tissue inhibitor of metalloproteinase 1 nor immunoprecipitated by antisera to MMPs, suggesting that the 97-kDa protease is not a MMP. Together, these data suggest that MMPs along with an unidentified 97-kDa protease degrade IGFBP-5 in MC3T3-E1 cell cultures. Because truncated forms of IGFBP-5 have been shown to enhance the action of IGF in bone cells, IGFBP-5 proteases may be instrumental in IGF-mediated bone morphogenesis.
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PMID:Characterization of insulin-like growth factor-binding protein 5-degrading proteases produced throughout murine osteoblast differentiation. 754 45

In the pig, the corpus luteum (CL) can develop and function autonomous of pituitary gonadotropins for approximately 12 days. We hypothesized that the insulin-like growth factor (IGF) system may play an autocrine/paracrine luteotrophic role(s) during this period. In this study, we monitored the expression (i.e., steady-state levels of mRNAs) of IGF-I and IGF binding proteins (IGFBP)-2, -3, -4, -5, and -6 mRNAs in whole CL and in small and large luteal cells on Days 4-16 of the estrous cycle. CL were dissociated with collagenase, and large and small luteal cells were isolated by centrifugal elutriation. Whole CL and luteal cells were extracted to isolate total or poly(A)+ RNA, which was subjected to Northern and/or dot-blot analyses using [32P]-labeled cDNA probes for IGF-I and IGFBP-2, -3, -4, -5, and -6. Northern blots showed readily detectable transcripts for IGF-I (6.7 and 0.9 kb), IGFBP-2 (1.8 kb), IGFBP-3 (2.8 kb), IGFBP-4 (2.6 kb), and IGFBP-5 (6.0 kb), but not for IGFBP-6. IGFBP-3 and -5 transcripts were observed mainly in small luteal cells, while IGFBP-2 and -4 were seen in both cell types. Dot-blot analyses for IGF-I and IGFBP-3 mRNAs were performed on total RNA from small and large luteal cells; blots were counter-probed with 3-phosphoglyceraldehyde dehydrogenase (p-GAD) cDNA to assess RNA quantity and quality. IGF-I mRNA (ratio IGF-I:p-GAD mRNA) expression was approximately 2-fold greater in small than in large luteal cells on Days 4-10. However, steady-state levels of IGF-I mRNA in small, but not large, luteal cells decreased significantly on Days 12-16 (vs. Days 4-10). IGFBP-3 mRNA expression was significantly greater (approximately 3-fold) in small than in large luteal cells but did not vary significantly between Days 4-10 and 12-16 for either cell type. We conclude that porcine CL express mRNAs for IGF-I and IGFBP-2, -3, -4, and -5, and that while small luteal cells are the major sources of IGF-I and IGFBP-3 and -5, IGFBP-2 and -4 appear to be expressed to approximately the same extent in small and large luteal cells. These results further suggest that the IGF-I/IGF system may have autocrine/paracrine regulatory actions in CL development/function in the pig.
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PMID:Expression of the messenger ribonucleic acids for insulin-like growth factor-I and insulin-like growth factor binding proteins in porcine corpora lutea. 878 84

To identify proteins that are lost during the establishment of the transformed phenotype of a tumor cell, we have prepared a subtracted cDNA library with mRNA from normal human fibroblasts and from their matched SV40 transformed counterparts. More than 40 clones were obtained that showed a dramatic reduction in their relative expression after oncogenic transformation. The proteins encoded by these clones could be grouped into four distinct classes: extracellular matrix proteins (fibronectin, beta ig-h3, collagen VI), enzymes (collagenase, urokinase), cytoskeletal proteins (vinculin, SM22) and regulatory proteins (beta-glycan, integrin-associated protein, myosin kinase, IGFBP-5). Six novel gene products were discovered during these experiments, including a novel serine protease, a zyxin-like protein, an ankyrin-like protein and a GTP-binding protein. Only four of all the transformation-sensitive cDNAs were consistently down-regulated when a variety of cell lines derived from spontaneous mesenchymal tumors was investigated: beta ig-h3, collagen VI, the novel ankyrin-like protein, and IGFBP-5. It is likely that these gene products play an important role in the maintenance of the normal phenotype.
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PMID:Down-regulated proteins of mesenchymal tumor cells. 951 34

Insulin-like growth factor binding proteins (IGFBP) proteases have been proposed to be involved in changes of serum IGFBP pattern during pregnancy. IGFBP-4 and -5 are degraded specifically by proteases in pregnancy serum in vitro, whereas IGFBP-3 proteolytic activity was also detected in nonpregnancy serum. To identify and characterize IGFBP proteases, human pregnancy serum was fractionated by size exclusion chromatography revealing IGFBP-4 protease activities in fractions coeluting with proteins of approximately 600-kDa and 50- to 100-kDa molecular mass. In both fractions, a predominant 50-kDa gelatinase was found, suggesting that parts of the gelatinase activity might aggregate or are complexed with other proteins forming a higher molecular complex. Hydroxyapatite chromatography and chromatofocusing of the 50- to 100-kDa serum fraction showed that the IGFBP-4 protease and the 50-kDa gelatinase activity were copurified. When the 50-kDa gelatinase-containing band was excised from the polyacrylamide gel, it exhibited IGFBP-4 proteolytic activity, resulting in the formation of 17- and 10-kDa fragments. [125I] IGFBP substrate zymography combined with fragment blotting showed that the 1,10-phenanthroline-sensitive 50-kDa protease activity purified by chromatofocusing also cleaved IGFBP-3 and -5. Other proteases detected in pregnancy serum fractions with Mr estimates of 79-, 30-, and 22-kDa degraded IGFBP-3 and -5 but not IGFBP-4. [125I] IGFBP-5 substrate zymography revealed that the 30-kDa IGFBP protease was inhibited by serine protease inhibitors. Whereas 1,10-phenanthroline inhibited the IGFBP proteolytic activity in the solution assay, serine protease inhibitors failed to affect proteolysis, indicating the predominant contribution of the metalloproteinase to IGFBP proteolysis. Tissue inhibitors of matrix metalloproteinases-1 and -2 revealed weak or no inhibition of IGFBP-4 and -5 proteolytic activity, whereas a hydroxamic acid-based inhibitor, potentially inhibiting disintegrin metalloproteases, completely prevented the proteolysis of IGFBPs. Whereas no specific immunoreactivity of the 50-kDa protein with antimatrix metalloproteinase-1, -2, -3, -9, or -13 antibodies was observed, antidisintegrin domain-specific antibodies bound to the 50-kDa gelatinase. These studies provide the first direct biochemical evidence that human pregnancy serum contains a 50-kDa IGFBP protease with properties of a soluble disintegrin metalloproteinase that appears to be potentially involved in regulating IGF bioavailability for placental and fetal growth.
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PMID:Proteolysis of insulin-like growth factor binding proteins by a novel 50-kilodalton metalloproteinase in human pregnancy serum. 952 34