Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new member of the membrane-type matrix metalloproteinase (MT-MMP) subfamily tentatively named MT5-MMP was isolated from mouse brain cDNA library. It is predicted to contain (i) a candidate signal sequence, (ii) a propeptide region with the highly conserved PRCGVPD sequence, (iii) a potential furin recognition motif RRRRNKR, (iv) a zinc-binding catalytic domain, (v) a hemopexin-like domain, (vi) a 24-residue hydrophobic domain as a potential transmembrane domain, and (vii) a short cytosolic domain. Reverse transcriptase-polymerase chain reaction analysis of its transcripts indicates that MT5-MMP is expressed in a brain-specific manner consistent with the origin of its EST clone from cerebellum. It is also highly expressed during embryonic development at stages day 11 and 15. Like other MT-MMPs, MT5-MMP specifically activates progelatinase A when co-expressed in Madin-Darby canine kidney cells. Its ability to activate progelatinase A is dependent on its proteolytic activity since a mutation converting Glu to Ala in the zinc binding motif HE255LGH renders MT5-MMP inactive against progelatinase A. In contrast to other MT-MMPs, MT5-MMP tends to shed from cell surface as soluble proteinases, thus offering flexibility as both a cell bound and soluble proteinase for extracellular matrix remodeling processes. Taken together, these properties serve to distinguish MT5-MMP as a versatile MT-MMP playing an important role in extracellular matrix remodeling events in the brain and during embryonic development.
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PMID:Identification and characterization of the fifth membrane-type matrix metalloproteinase MT5-MMP. 1008 37

We investigated the gene expression and intracellular activity of processing protease furin and its involvement in the process of membrane type 1-matrix metalloproteinase (MT1-MMP) activation in rabbit dermal fibroblasts. When the rabbit fibroblasts were treated with concanavalin A (ConA), pro-MMP-2 was converted to an active 62-kDa MMP-2 through the appearance of a 64-kDa intermediate MMP-2. The ConA-induced pro-MMP-2 activation resulted from increasing the gene expression and production of MT1-MMP in the rabbit fibroblasts. Reverse transcriptase-polymerase chain reaction demonstrated that in rabbit dermal fibroblasts furin mRNA was detected and, unlike MT1-MMP, was not increased by ConA. These findings are further supported by the fact that the intracellular furin activity also was constitutively detected and was unchanged by the ConA treatment. Very similar phenomena were also observed in human uterine cervical fibroblasts, which are known to produce MT1-MMP by ConA stimulation. These results suggest that the expression of the furin gene and the intracellular activity are not regulated by ConA. On the other hand, neither a synthetic furin inhibitor, decanoyl-RVKR-CH(2)Cl (25-100 microM) nor a furin antisense oligonucleotide (40 microM) inhibited the MT1-MMP-mediated pro-MMP-2 activation in ConA-treated rabbit dermal fibroblasts, whereas these compounds interfered with pro-MMP-2 activation in ConA-treated human uterine cervical fibroblasts. Nonetheless, the furin antisense oligonucleotide completely suppressed furin gene expression in both rabbit and human fibroblasts. These results suggest that furin does not participate in the process of MT1-MMP activation induced by ConA in rabbit dermal fibroblasts.
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PMID:Furin-independent pathway of membrane type 1-matrix metalloproteinase activation in rabbit dermal fibroblasts. 1060 Dec 93

LR7/8B and ApoER2 are recently discovered members of the low density lipoprotein (LDL) receptor family. Although structurally different, these two proteins are derived from homologous genes in chicken and man by alternative splicing and contain 7 or 8 LDL receptor ligand-binding repeats. Here we present the cDNA for ApoER2 cloned from mouse brain and describe splice variants in the ligand binding domain of this protein, which are distinct from those present in man and chicken. The cloned cDNA is coding for a receptor with only five LDL receptor ligand-binding repeats, i.e. comprising repeats 1-3, 7, and 8. Reverse transcriptase-polymerase chain reaction analysis of mRNA from murine brain revealed the existence of two additional transcripts. One is lacking repeat 8, and in the other repeat 8 is substituted for by a 13-amino acid insertion with a consensus site for furin cleavage arising from an additional small exon present in the murine gene. None of the transcripts in the mouse, however, contain repeats 4-6. In murine placenta only the form containing repeats 1-3 and 7 and the furin cleavage site is detectable. Analysis of the corresponding region of the murine gene showed the existence of 6 exons coding for a total of 8 ligand binding repeats, with one exon encoding repeats 4-6. Exon trapping experiments demonstrated that this exon is constitutively spliced out in all murine transcripts. Thus, the murine ApoER2 gene codes for receptor variants harboring either 4 or 5 binding repeats only. Recombinant expression of the 5-repeat and 4-repeat variants showed that repeats 1-3, 7, and 8 are sufficient for binding of beta-very low density lipoprotein and reelin, but not for recognition of alpha(2)-macroglobulin, which binds to the avian homologue of ApoER2 harboring 8 ligand binding repeats.
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PMID:Alternative splicing in the ligand binding domain of mouse ApoE receptor-2 produces receptor variants binding reelin but not alpha 2-macroglobulin. 1129 45

Cell therapy may have the potential for the treatment of Type I diabetes. To date, cells suitable for this purpose have not been developed. This study investigates the feasibility of modifying Vero, a cell line that may be considered safe to implant into humans, for this purpose. Stable Vero transfectants containing full-length human preproinsulin complementary deoxyribonucleic acid (cDNA) were generated using a liposomal transfection reagent. Reverse transcriptase-polymerase chain reaction, immunocytochemistry, Western blotting, and enzyme-linked immunosorbent assays were used to assess the resulting cells. Proinsulin was expressed but was not processed to insulin by these cells. Proinsulin cDNA was genetically modified, resulting in a form that is furin sensitive. The resulting stably transfected Vero clones constitutively release approximately 34%/h (32.68 +/- 2.21 to 35.62 +/- 3.14%) of the product formed, approximately 62% (59.99 +/- 6.45 to 64.64 +/- 4.57%) of which is mature insulin. These Vero transfectants did not exhibit glucose-stimulated insulin secretion. As GLUT2 and glucokinase (GCK) are not constitutively expressed by these cells, human GLUT2 cDNA and GCK cDNA were cotransfected with furin-sensitive preproinsulin cDNA into Vero cells. Insulin and GCK proteins were detected in the cytoplasmic region of the resulting cells, whereas GLUT2 was predominantly expressed in the nucleus. Coexpression of GLUT2 and GCK did not result in glucose-stimulated insulin secretion. The results from this study demonstrate the feasibility of engineering a relatively "safe" nonbeta cell line to produce human insulin. Coexpression of GLUT2 and GCK, at the levels achieved here, is not adequate enough to induce glucose-stimulated insulin secretion in such cells; the subcellular location of transfected components may be relevant.
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PMID:Engineering Vero cells to secrete human insulin. 1202 63

The structure and distribution of PC5-A, a prohormone convertase that is thought to be involved in post-translational processing of peptide hormone and neuropeptide precursors, have not been investigated in submammalian vertebrates. In the present study, we characterized the cDNA encoding PC5-A in the European green frog Rana esculenta. The frog PC5-A cDNA encodes a 913-amino acid protein that encompasses a 28-amino acid signal peptide, the Asp/His/Ser catalytic triad found in all serine proteinases of the subtilisin family, and two potential N-linked glycosylation sites located in a C-terminal cysteine-rich domain. Reverse transcriptase polymerase chain reaction amplification showed that PC5-A mRNA is expressed in various organs including the brain, spinal cord, pituitary, lung, liver, intestine, and testis, but not in the stomach and pancreas. The distribution of PC5-A mRNA in the frog brain was studied by in situ hybridization histochemistry. Intense expression was observed in the mitral cellular layer of the olfactory bulb, the nucleus of the diagonal band of Broca, the anterior preoptic area, and the suprachiasmatic and ventral hypothalamic nuclei. The expression pattern of PC5-A mRNA in the central nervous system of anuran amphibians was consistent with the implication of this prohormone convertase in the processing of various neuropeptide precursors.
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PMID:Molecular characterization of the cDNA and localization of the mRNA encoding the prohormone convertase PC5-A in the European green frog. 1250 14

A novel common integration site for the mouse mammary tumor virus (MMTV) was identified (designated Int7) in five independently arising mouse mammary tumors. The insertion sites all cluster within a 1-kb region that is 2 to 3 kb 5' of the transcription initiation site of a gene, 2610028F08RIK, whose gene product contains furin-like and thrombospondin-like sequences. Expression of Int7 is normally very low or silent during various stages of mammary gland development, but MMTV integration at this site results in the activation of high steady-state levels of expression of the gene. These five tumors were also found to have two or three additional viral insertions, which in each case occurred flanking a member of either the Wnt and/or FGF gene family. Reverse transcriptase PCR results demonstrated that each of the viral insertions led to elevated expression of the presumed target flanking genes.
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PMID:A new common integration site, Int7, for the mouse mammary tumor virus in mouse mammary tumors identifies a gene whose product has furin-like and thrombospondin-like sequences. 1601 73

Human mast cells have been shown to release histamine in response to the neuropeptide alpha-melanocyte-stimulating hormone (alpha-MSH), but it is unknown whether these cells express proopiomelanocortin (POMC) or POMC-derived peptides. We therefore examined highly purified human skin mast cells and a leukemic mast cell line-1 (HMC-1) for their ability to express POMC and members of the prohormone convertase (PC) family known to process POMC. Furthermore, we investigated whether these cells store and secrete alpha-MSH. Reverse transcriptase-PCR (RT-PCR) analysis revealed that both skin mast cells and HMC-1 cells express POMC mRNA and protein. Expression of the POMC gene at the RNA level in HMC-1 cells could be confirmed by Northern blotting. Transcripts for both PC1 and furin convertase were detectable in skin-derived mast cells and HMC-1 cells, as shown by RT-PCR. In contrast, PC2 transcripts were detected only in skin mast cells, whereas transcripts for paired basic amino acid converting enzyme 4 (PACE4) were present only in HMC-1 cells. Radioimmunoassays performed on cell lysates and cell culture supernatants from human skin-derived mast cells disclosed immunoreactive amounts of alpha-MSH in both fractions. Stimulation with an anti-IgE antibody significantly reduced intracellular alpha-MSH and increased extracellular levels, indicating IgE-mediated secretion of this neuropeptide. Our findings show that human mast cells are active players in the cutaneous POMC system. Mast cell-derived alpha-MSH may contribute to cutaneous hyperpigmentation as seen in patients with urticaria pigmentosa. Moreover, IgE-dependent release of alpha-MSH suggests an immunomodulatory role of this neurohormone during inflammatory and allergic reactions of the skin.
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PMID:Human mast cells in the neurohormonal network: expression of POMC, detection of precursor proteases, and evidence for IgE-dependent secretion of alpha-MSH. 1691 90

The apicomplexan parasite Cryptosporidium is a significant cause of diarrheal disease worldwide. Previously, we reported that a Cryptosporidium parvum subtilisin-like serine protease activity with furin-type specificity cleaves gp40/15, a glycoprotein that is proteolytically processed into gp40 and gp15, which are implicated in mediating infection of host cells. Neither the enzyme(s) responsible for the protease activity in C. parvum lysates nor those that process gp40/15 are known. There are no furin or other proprotein convertase genes in the C. parvum genome. However, a gene encoding CpSUB1, a subtilisin-like serine protease, is present. In this study, we cloned the CpSUB1 genomic sequence and expressed and purified the recombinant prodomain. Reverse transcriptase PCR analysis of RNA from C. parvum-infected HCT-8 cells revealed that CpSUB1 is expressed throughout infection in vitro. In immunoblots, antiserum to the recombinant CpSUB1 prodomain revealed two major bands, of approximately 64 kDa and approximately 48 kDa, for C. parvum lysates and proteins "shed" during excystation. In immunofluorescence assays, the antiserum reacted with the apical region of sporozoites and merozoites. The recombinant prodomain inhibited protease activity and processing of recombinant gp40/15 by C. parvum lysates but not by furin. Since prodomains are often selective inhibitors of their cognate enzymes, these results suggest that CpSUB1 may be a likely candidate for the protease activity in C. parvum and for processing of gp40/15. Importantly, the recombinant prodomain inhibited C. parvum infection of HCT-8 cells. These studies indicate that CpSUB1 plays a significant role in infection of host cells by the parasite and suggest that this enzyme may serve as a target for intervention.
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PMID:Role of CpSUB1, a subtilisin-like protease, in Cryptosporidium parvum infection in vitro. 1916 60

Recently, a mini-review was published in the Medical Hypotheses journal by Usul Afsar entitled 2019-nCoV-SARS-CoV-2 (COVID-19) infection: Cruciality of Furin and relevance with cancer. Previous studies have pointed out that disruption of the proteolytic cleavage of proteins can promote infectious and non-infectious diseases. The last few weeks have been marked by an important revelation concerning the pathophysiology of SARS-CoV-2. This new coronavirus disease (COVID-19) is a highly contagious and transmissible acute respiratory infectious disorder. SARS-CoV-2 is composed of RNA-dependent RNA polymerase and structural proteins including Spike protein (S protein). Interestingly, the FURIN, one of the proproteins of the convertase family, plays a crucial role in the maturation of viral glycoproteins. In addition, many viruses including coronaviruses, exploit FURIN for the activation of their glycoproteins. Recent data indicate that SARS-CoV-2 enters human cells by binding to angiotensin-converting enzyme 2. Subsequently, the S protein is cleaved by transmembrane protease serine 2 with the help of FURIN which facilitates the entry of the virus into the cell after binding. Furthermore, it seems that FURIN is implicated in the pathogenesis of SARS-CoV-2 and potentially in the increased rates of human-to-human transmission.
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PMID:Expression of angiotensin-converting enzyme 2 and proteases in COVID-19 patients: A potential role of cellular FURIN in the pathogenesis of SARS-CoV-2. 3251 90