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1.
Artigo em Inglês | MEDLINE | ID: mdl-31482488

RESUMO

A crucial step for tumor cell extravasation and metastasis is the migration through the extracellular matrix, which requires proteolytic activity. Hence, proteases, particularly matrix metalloproteases (MMPs), have been discussed as therapeutic targets and their inhibition should diminish tumor growth and metastasis. The metalloproteases meprin α and meprin ß are highly abundant on intestinal enterocytes and their expression was associated with different stages of colorectal cancer. Due to their ability to cleave extracellular matrix (ECM) components, they were suggested as pro-tumorigenic enzymes. Additionally, both meprins were shown to have pro-inflammatory activity by cleaving cytokines and their receptors, which correlates with chronic intestinal inflammation and associated conditions. On the other hand, meprin ß was identified as an essential enzyme for the detachment and renewal of the intestinal mucus, important to prevent bacterial overgrowth and infection. Considering this, it is hard to estimate whether high activity of meprins is generally detrimental or if these enzymes have also protective functions in certain cancer types. For instance, for colorectal cancer, patients with high meprin ß expression in tumor tissue exhibit a better survival prognosis, which is completely different to prostate cancer. This demonstrates that the very same enzyme may have contrary effects on tumor initiation and growth, depending on its tissue and subcellular localization. Hence, precise knowledge about proteolytic enzymes is required to design the most efficient therapeutic options for cancer treatment. In this review, we summarize the current findings on meprins' functions, expression, and cancer-associated variants with possible implications for tumor progression and metastasis.

2.
Brain Behav Immun ; 2019 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-31401302

RESUMO

Alzheimer's disease (AD) is the most commonly diagnosed dementia but its underlying pathological mechanisms still unclear. Neuroinflammation and secretion of cytokines such as interleukin-6 (IL-6) accompany the main hallmarks of the disease: amyloid plaques and neurofibrillary tangles. In this study, we analyzed the role of IL-6 trans-signaling in two mouse models of AD, Tg2576 and 3xTg-AD mice. The inhibition of IL-6 trans-signaling partially rescued the AD-induced mortality in females of both models. Before amyloid plaques deposition, it reversed AD-induced changes in exploration and anxiety (but did not affect locomotion) in Tg2576 female mice. However, after plaque deposition the only behavioral trait affected by the inhibition of IL-6 trans-signaling was locomotion. Results in the Morris water maze suggest that cognitive flexibility was reduced by the blocking of the IL-6 trans-signaling in young and old Tg2576 female mice. The inhibition of IL-6 trans-signaling also decreased amyloid plaque burden in cortex and hippocampus, and Aß40 and Aß42 levels in the cortex, of Tg2576 female mice. The aforementioned changes might be correlated with changes in blood vessels and matrix structure and organization rather than changes in neuroinflammation. 3xTgAD mice showed a very mild phenotype regarding amyloid cascade, but results were in accordance with those of Tg2576 mice. These results strongly suggest that the inhibition of the IL-6 trans-signaling could represent a powerful therapeutic target in AD.

3.
FASEB J ; : fj201801371R, 2019 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-31381863

RESUMO

Meprin ß is a membrane-bound metalloprotease involved in extracellular matrix assembly and inflammatory processes in health and disease. A disintegrin and metalloproteinase (ADAM)10 and ADAM17 are physiologic relevant sheddases of inactive promeprin ß, which influences its substrate repertoire and subsequent biologic functions. Proteomic analysis also revealed several ADAMs as putative meprin ß substrates. Here, we demonstrate specific N-terminal processing of ADAM9, 10, and 17 by meprin ß and identify cleavage sites within their prodomains. Because ADAM prodomains can act as specific inhibitors, we postulate a role for meprin ß in the regulation of ADAM activities. Indeed, prodomain cleavage by meprin ß caused increased ADAM protease activities, as observed by peptide-based cleavage assays and demonstrated by increased ectodomain shedding activity. Direct interaction of meprin ß and ADAM proteases could be shown by immunofluorescence microscopy and immunoprecipitation experiments. As demonstrated by a bacterial activator of meprin ß and additional measurement of TNF-α shedding on bone marrow-derived macrophages, meprin ß/ADAM protease interactions likely influence inflammatory conditions. Thus, we identified a novel proteolytic pathway of meprin ß with ADAM proteases to control protease activities at the cell surface as part of the protease web.-Wichert, R., Scharfenberg, F., Colmorgen, C., Koudelka, T., Schwarz, J., Wetzel, S., Potempa, B., Potempa, J., Bartsch, J. W., Sagi, I., Tholey, A., Saftig, P., Rose-John, S., Becker-Pauly, C. Meprin ß induces activities of A disintegrin and metalloproteinases 9, 10, and 17 by specific prodomain cleavage.

4.
Int J Mol Sci ; 20(15)2019 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-31357561

RESUMO

Interleukin-11 (IL-11) has been associated with inflammatory conditions, bone homeostasis, hematopoiesis, and fertility. So far, these functions have been linked to classical IL-11 signaling via the membrane bound receptor (IL-11R). However, a signaling cascade via the soluble IL-11R (sIL-11R), generated by proteolytic cleavage, can also be induced. This process is called IL-11 trans-signaling. A disintegrin and metalloprotease 10 (ADAM10) and neutrophil elastase were described as ectodomain sheddases of the IL-11R, thereby inducing trans-signaling. Furthermore, previous studies employing approaches for the stimulation and inhibition of endogenous ADAM-proteases indicated that ADAM10, but not ADAM17, can cleave the IL-11R. Herein, we show that several metalloproteases, namely ADAM9, ADAM10, ADAM17, meprin ß, and membrane-type 1 matrix metalloprotease/matrix metalloprotease-14 (MT1-MMP/MMP-14) when overexpressed are able to shed the IL-11R. All sIL-11R ectodomains were biologically active and capable of inducing signal transducer and activator of transcription 3 (STAT3) phosphorylation in target cells. The difference observed for ADAM10/17 specificity compared to previous studies can be explained by the different approaches used, such as stimulation of protease activity or making use of cells with genetically deleted enzymes.

5.
Cell Mol Life Sci ; 76(16): 3193-3206, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31201463

RESUMO

Alzheimer's Disease (AD) is the sixth-leading cause of death in industrialized countries. Neurotoxic amyloid-ß (Aß) plaques are one of the pathological hallmarks in AD patient brains. Aß accumulates in the brain upon sequential, proteolytic processing of the amyloid precursor protein (APP) by ß- and γ-secretases. However, so far disease-modifying drugs targeting ß- and γ-secretase pathways seeking a decrease in the production of toxic Aß peptides have failed in clinics. It has been demonstrated that the metalloproteinase meprin ß acts as an alternative ß-secretase, capable of generating truncated Aß2-x peptides that have been described to be increased in AD patients. This indicates an important ß-site cleaving enzyme 1 (BACE-1)-independent contribution of the metalloprotease meprin ß within the amyloidogenic pathway and may lead to novel drug targeting avenues. However, meprin ß itself is embedded in a complex regulatory network. Remarkably, the anti-amyloidogenic α-secretase a disintegrin and metalloproteinase domain-containing protein 10 (ADAM10) is a direct competitor for APP at the cell surface, but also a sheddase of inactive pro-meprin ß. Overall, we highlight the current cellular, molecular and structural understanding of meprin ß as alternative ß-secretase within the complex protease web, regulating APP processing in health and disease.


Assuntos
Proteína ADAM10/metabolismo , Metaloendopeptidases/metabolismo , Proteína ADAM10/química , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Metaloendopeptidases/química , Presenilina-1/metabolismo , Proteólise , Serina Endopeptidases/metabolismo
6.
Cell Mol Life Sci ; 2019 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-31209506

RESUMO

Disintegrin and metalloproteinases (ADAMs) 10 and 17 can release the extracellular part of a variety of membrane-bound proteins via ectodomain shedding important for many biological functions. So far, substrate identification focused exclusively on membrane-anchored ADAM10 and ADAM17. However, besides known shedding of ADAM10, we identified ADAM8 as a protease capable of releasing the ADAM17 ectodomain. Therefore, we investigated whether the soluble ectodomains of ADAM10/17 (sADAM10/17) exhibit an altered substrate spectrum compared to their membrane-bound counterparts. A mass spectrometry-based N-terminomics approach identified 134 protein cleavage events in total and 45 common substrates for sADAM10/17 within the secretome of murine cardiomyocytes. Analysis of these cleavage sites confirmed previously identified amino acid preferences. Further in vitro studies verified fibronectin, cystatin C, sN-cadherin, PCPE-1 as well as sAPP as direct substrates of sADAM10 and/or sADAM17. Overall, we present the first degradome study for sADAM10/17, thereby introducing a new mode of proteolytic activity within the protease web.

7.
J Cell Sci ; 132(11)2019 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-31076514

RESUMO

The extracellular metalloprotease meprin ß is expressed as a homodimer and is primarily membrane bound. Meprin ß can be released from the cell surface by its known sheddases ADAM10 and ADAM17. Activation of pro-meprin ß at the cell surface prevents its shedding, thereby stabilizing its proteolytic activity at the plasma membrane. We show that a single amino acid exchange variant (G32R) of meprin ß, identified in endometrium cancer, is more active against a peptide substrate and the IL-6 receptor than wild-type meprin ß. We demonstrate that the change to an arginine residue at position 32 represents an additional activation site used by furin-like proteases in the Golgi, which consequently leads to reduced shedding by ADAM17. We investigated this meprin ß G32R variant to assess cell proliferation, invasion through a collagen IV matrix and outgrowth from tumor spheroids. We found that increased meprin ß G32R activity at the cell surface reduces cell proliferation, but increases cell invasion.

8.
FASEB J ; 33(6): 7490-7504, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30916990

RESUMO

Biologic activity of proteases is mainly characterized by the substrate specificity, tissue distribution, and cellular localization. The human metalloproteases meprin α and meprin ß share 41% sequence identity and exhibit a similar cleavage specificity with a preference for negatively charged amino acids. However, shedding of meprin α by furin on the secretory pathway makes it a secreted enzyme in comparison with the membrane-bound meprin ß. In this study, we identified human meprin α and meprin ß as forming covalently linked membrane-tethered heterodimers in the early endoplasmic reticulum, thereby preventing furin-mediated secretion of meprin α. Within this newly formed enzyme complex, meprin α was able to be activated on the cell surface and detected by cleavage of a novel specific fluorogenic peptide substrate. However, the known meprin ß substrates amyloid precursor protein and CD99 were not shed by membrane-tethered meprin α. On the other hand, being linked to meprin α, activation of or substrate cleavage by meprin ß on the cell surface was not altered. Interestingly, proteolytic activity of both proteases was increased in the heteromeric complex, indicating an increased proteolytic potential at the plasma membrane. Because meprins are susceptibility genes for inflammatory bowel disease (IBD), and to investigate the physiologic impact of the enzyme complex, we performed transcriptome analyses of intestinal mucosa from meprin-knockout mice. Comparison of the transcriptional gene analysis data with gene analyses of IBD patients revealed that different gene subsets were dysregulated if meprin α was expressed alone or in the enzyme complex, demonstrating the physiologic and pathophysiological relevance of the meprin heterodimer formation.-Peters, F., Scharfenberg, F., Colmorgen, C., Armbrust, F., Wichert, R., Arnold, P., Potempa, B., Potempa, J., Pietrzik, C. U., Häsler, R., Rosenstiel, P., Becker-Pauly, C. Tethering soluble meprin α in an enzyme complex to the cell surface affects IBD-associated genes.

9.
Sci Rep ; 9(1): 546, 2019 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-30679641

RESUMO

Vertebrate fetuins are multi-domain plasma-proteins of the cystatin-superfamily. Human fetuin-A is also known as AHSG, α2-Heremans-Schmid-glycoprotein. Gene-knockout in mice identified fetuin-A as essential for calcified-matrix-metabolism and bone-mineralization. Fetuin-B deficient mice, on the other hand, are female infertile due to zona pellucida 'hardening' caused by the metalloproteinase ovastacin in unfertilized oocytes. In wildtype mice fetuin-B inhibits the activity of ovastacin thus maintaining oocytes fertilizable. Here we asked, if fetuins affect further proteases as might be expected from their evolutionary relation to single-domain-cystatins, known as proteinase-inhibitors. We show that fetuin-A is not an inhibitor of any tested protease. In stark contrast, the closely related fetuin-B selectively inhibits astacin-metalloproteinases such as meprins and ovastacin, but not astacins of the tolloid-subfamily, nor any other proteinase. The analysis of fetuin-B expressed in various mammalian cell types, insect cells, and truncated fish-fetuin expressed in bacteria, showed that the cystatin-like domains alone are necessary and sufficient for inhibition. This report highlights fetuin-B as a specific antagonist of ovastacin and meprin-metalloproteinases. Control of ovastacin was shown to be indispensable for female fertility. Meprin inhibition, on the other hand, renders fetuin-B a potential key-player in proteolytic networks controlling angiogenesis, immune-defense, extracellular-matrix-assembly and general cell-signaling, with implications for inflammation, fibrosis, neurodegenerative disorders and cancer.

10.
FEBS Open Bio ; 8(12): 2011-2021, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30524951

RESUMO

BMP-1/tolloid-like proteinases belong to the astacin family of human metalloproteinases, together with meprins and ovastacin. They represent promising targets to treat or prevent a wide range of diseases such as fibrotic disorders or cancer. However, the study of their pathophysiological roles is still impaired by the lack of well-characterized inhibitors and the questions that remain regarding their selectivity and in vivo efficiency. As a first step towards the identification of suitable tools to be used in functional studies, we have undertaken a systematic comparison of seven molecules known to affect the proteolytic activity of human astacins including three hydroxamates (FG-2575, UK383,367, S33A), the protein sizzled, a new phosphinic inhibitor (RXP-1001) and broad-spectrum protease inhibitors (GM6001, actinonin). Their efficacy in vitro, their cellular toxicity and efficacy in cell cultures were thoroughly characterized. We found that these molecules display very different potency and selectivity profiles, with hydroxamate FG-2575 and the protein sizzled being very powerful and selective inhibitors of BMP-1, whereas phosphinic peptide RXP-1001 behaves as a broad-spectrum inhibitor of astacins. Their use should therefore be carefully considered in agreement with the aim of the study to avoid result misinterpretation.

11.
FEBS J ; 2018 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-30422384

RESUMO

Matrix metalloproteinases (MMPs) are secreted as proenzymes, containing propeptides that interact with the catalytic zinc, thereby controlling MMP activation. The MMP-9 propeptide is unique in the MMP family because of its post-translational modification with an N-linked oligosaccharide. ProMMP-9 activation by MMP-3 occurs stepwise by cleavage of the propeptide in an aminoterminal (pro-AT) and carboxyterminal (pro-CT) peptide. We chemically synthesized aglycosyl pro-AT and pro-CT and purified recombinant glycosylated pro-ATS f-9 . First, we report new cleavage sites in the MMP-9 propeptide by MMP-3 and neutrophil elastase. Additionally, we demonstrated with the use of western blot analysis a higher resistance of glycosylated versus aglycosyl pro-AT against proteolysis by MMP-3, MMP-9, meprin α, neutrophil elastase and by protease-rich synovial fluids from rheumatoid arthritis patients. Moreover, we investigated the effect of glycosylation on proteolytic activation of human proMMP-9 with the use of zymography and dye-quenched gelatin cleavage analysis. Compared to recombinant Sf-9 proMMP-9 glycoforms, larger oligosaccharides of human neutrophil proMMP-9 increased resistance against proteolytic activation. Additionally, proMMP-9 from Congenital Disorder of Glycosylation patients, compared to healthy controls, showed a higher activation rate by MMP-3. Finally, we demonstrated that glycan-galectin-3 interactions reduced proMMP-9 activation. In conclusion, modification of MMP-9 propeptide glycosylation is a fine-tuning mechanism and co-determines the specific activity of MMP-9 in physiology and pathology. ENZYMES: MMP-9 EC 3.4.24.35, MMP-3 EC 3.4.24.17, meprin α EC 3.4.24.18, neutrophil elastase EC 3.4.21.37, trypsin EC 3.4.21.4 and PNGase F EC 3.5.1.52.

12.
Acta Neuropathol ; 2018 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-30426203

RESUMO

Brain accumulation and aggregation of amyloid-ß (Aß) peptides is a critical step in the pathogenesis of Alzheimer's disease (AD). Full-length Aß peptides (mainly Aß1-40 and Aß1-42) are produced through sequential proteolytic cleavage of the amyloid precursor protein (APP) by ß- and γ-secretases. However, studies of autopsy brain samples from AD patients have demonstrated that a large fraction of insoluble Aß peptides are truncated at the N-terminus, with Aß4-x peptides being particularly abundant. Aß4-x peptides are highly aggregation prone, but their origin and any proteases involved in their generation are unknown. We have identified a recognition site for the secreted metalloprotease ADAMTS4 (a disintegrin and metalloproteinase with thrombospondin motifs 4) in the Aß peptide sequence, which facilitates Aß4-x peptide generation. Inducible overexpression of ADAMTS4 in HEK293 cells resulted in the secretion of Aß4-40 but unchanged levels of Aß1-x peptides. In the 5xFAD mouse model of amyloidosis, Aß4-x peptides were present not only in amyloid plaque cores and vessel walls, but also in white matter structures co-localized with axonal APP. In the ADAMTS4-/- knockout background, Aß4-40 levels were reduced confirming a pivotal role of ADAMTS4 in vivo. Surprisingly, in the adult murine brain, ADAMTS4 was exclusively expressed in oligodendrocytes. Cultured oligodendrocytes secreted a variety of Aß species, but Aß4-40 peptides were absent in cultures derived from ADAMTS4-/- mice indicating that the enzyme was essential for Aß4-x production in this cell type. These findings establish an enzymatic mechanism for the generation of Aß4-x peptides. They further identify oligodendrocytes as a source of these highly amyloidogenic Aß peptides.

13.
Thromb Haemost ; 118(10): 1790-1802, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30235485

RESUMO

Pulmonary arterial hypertension (PAH) is a rare disease characterized by increased pulmonary pressure and vascular remodelling as a consequence of smooth muscle cell proliferation, endothelial cell dysfunction and inflammatory infiltrates. Meprin α is a metalloproteinase whose substrates include adhesion and cell-cell contact molecules involved in the process of immune cell extravasation. In this study, we aimed to unravel the role of meprin α in PAH-induced vascular remodelling. Our results showed that meprin α was present in the apical membrane of endothelial cells in the lungs and pulmonary arteries of donors and idiopathic PAH (IPAH) patients. Elevated circulating meprin α levels were detected in the plasma of IPAH patients. In vitro binding assays and electron microscopy confirmed binding of meprin α to the glycocalyx of human pulmonary artery endothelial cells (hPAECs). Enzymatic and genetic approaches identified heparan sulphate (HS) as an important determinant of the meprin α binding capacity to hPAEC. Meprin α treatment protected from excessive neutrophil infiltration and the protective effect observed in the presence of neutrophils was partially reversed by removal of HS from hPAEC. Importantly, HS levels in pulmonary arteries were decreased in IPAH patients and binding of meprin α to HS was impaired in IPAH hPAEC. In summary, our results suggest a role of HS in docking meprin α to the endothelium and thus in the modulation of inflammatory cell extravasation. In IPAH, the decreased endothelial HS results in the reduction of meprin α binding which might contribute to enhanced inflammatory cell extravasation and potentially to pathological vascular remodelling.

14.
J Exp Med ; 215(4): 1205-1225, 2018 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-29472497

RESUMO

Colorectal cancer is treated with antibodies blocking epidermal growth factor receptor (EGF-R), but therapeutic success is limited. EGF-R is stimulated by soluble ligands, which are derived from transmembrane precursors by ADAM17-mediated proteolytic cleavage. In mouse intestinal cancer models in the absence of ADAM17, tumorigenesis was almost completely inhibited, and the few remaining tumors were of low-grade dysplasia. RNA sequencing analysis demonstrated down-regulation of STAT3 and Wnt pathway components. Because EGF-R on myeloid cells, but not on intestinal epithelial cells, is required for intestinal cancer and because IL-6 is induced via EGF-R stimulation, we analyzed the role of IL-6 signaling. Tumor formation was equally impaired in IL-6-/- mice and sgp130Fc transgenic mice, in which only trans-signaling via soluble IL-6R is abrogated. ADAM17 is needed for EGF-R-mediated induction of IL-6 synthesis, which via IL-6 trans-signaling induces ß-catenin-dependent tumorigenesis. Our data reveal the possibility of a novel strategy for treatment of colorectal cancer that could circumvent intrinsic and acquired resistance to EGF-R blockade.

15.
Cell Rep ; 21(8): 2090-2103, 2017 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-29166602

RESUMO

The host metalloprotease meprin ß is required for mucin 2 (MUC2) cleavage, which drives intestinal mucus detachment and prevents bacterial overgrowth. To gain access to the cleavage site in MUC2, meprin ß must be proteolytically shed from epithelial cells. Hence, regulation of meprin ß shedding and activation is important for physiological and pathophysiological conditions. Here, we demonstrate that meprin ß activation and shedding are mutually exclusive events. Employing ex vivo small intestinal organoid and cell culture experiments, we found that ADAM-mediated shedding is restricted to the inactive pro-form of meprin ß and is completely inhibited upon its conversion to the active form at the cell surface. This strict regulation of meprin ß activity can be overridden by pathogens, as demonstrated for the bacterial protease Arg-gingipain (RgpB). This secreted cysteine protease potently converts membrane-bound meprin ß into its active form, impairing meprin ß shedding and its function as a mucus-detaching protease.


Assuntos
Adesinas Bacterianas/metabolismo , Cisteína Endopeptidases/metabolismo , Metaloendopeptidases/metabolismo , Metaloproteases/metabolismo , Sequência de Aminoácidos/genética , Animais , Membrana Celular/metabolismo , Células Epiteliais/metabolismo , Feminino , Células HEK293 , Humanos , Masculino , Metaloendopeptidases/genética , Camundongos Transgênicos , Mucina-2/genética , Mucina-2/metabolismo
16.
Oncotarget ; 8(33): 54873-54888, 2017 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-28903388

RESUMO

Transendothelial cell migration (TEM) is crucial for inflammation and metastasis. The adhesion molecule CD99 was shown to be important for correct immune cell extravasation and is highly expressed on certain cancer cells. Recently, we demonstrated that ectodomain shedding of CD99 by the metalloprotease meprin ß promotes TEM in vitro. In this study, we employed an acute inflammation model (air pouch/carrageenan) and found significantly less infiltrated cells in meprin ß knock-out animals validating the previously observed pro-inflammatory activity. To further analyze the impact of meprin ß on CD99 shedding with regard to cell adhesion and proliferation we characterized two lung cancer associated CD99 variants (D92H, D92Y), carrying point mutations at the main cleavage site. Interestingly, ectodomain shedding of these variants by meprin ß was still detectable. However the cleavage site shifted to adjacent positions. Nevertheless, expression of CD99 variants D92H and D92Y revealed partial misfolding and proteasomal degradation. A previously observed influence of CD99 on Src activation and increased proliferation could not be confirmed in this study, independent of wild-type CD99 or the variants D92H and D92Y. However, we identified meprin ß as a potent inducer of Src phosphorylation. Importantly, we found significantly increased cell migration when expressing the cancer-associated CD99 variant D92H compared to the wild-type protein.

17.
Biochim Biophys Acta Mol Cell Res ; 1864(11 Pt B): 2096-2104, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28502593

RESUMO

The zinc-endopeptidases meprin α and meprin ß are extracellular proteases involved in connective tissue homeostasis, intestinal barrier function and immunological processes. Meprins are unique among other extracellular proteases with regard to cleavage specificity and structure. Meprin α and meprin ß have a strong preference for negatively charged amino acids around the scissile bond, reflected by cleavage sites identified in procollagen I, the amyloid precursor protein (APP) and the interleukin-6 receptor (IL-6R). In this review we report on recent findings that summarize the complex molecular regulation of meprins, particular folding, activation and shedding. Dysregulation of meprin α and meprin ß is often associated with pathological conditions such as neurodegeneration, inflammatory bowel disease and fibrosis. Based on mouse models and patient data we suggest meprins as possible key regulators in the onset and progression of fibrotic disorders, leading to severe diseases such as pulmonary hypertension. This article is part of a Special Issue entitled: Proteolysis as a Regulatory Event in Pathophysiology edited by Stefan Rose-John.


Assuntos
Inflamação/genética , Metaloendopeptidases/genética , Proteólise , Sequência de Aminoácidos/genética , Animais , Regulação da Expressão Gênica/genética , Humanos , Inflamação/patologia , Metaloendopeptidases/biossíntese , Camundongos , Peptídeo Hidrolases/biossíntese , Peptídeo Hidrolases/genética
18.
Cell Calcium ; 65: 8-13, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28365001

RESUMO

The two metalloproteases meprin ß and bone morphogenetic protein 1 (BMP-1) are both members of the astacin protease family. They share specificity for negatively charged residues around the scissile bond and they are expressed in overlapping compartments of the human body. One important proteolytic substrate they share is pro-collagen I. Ablation of one of the two proteases however leads to different collagen I associated phenotypes in vivo. Over the last years calcium emerged as a regulator for the proteolytic activity of both enzymes. For meprin ß a reduction and for BMP-1 an increase in activity was reported under increasing calcium concentrations. Here we revisit different compartments that rely on pro-collagen I maturation and explore the crystal structure of both proteases to highlight possible calcium binding sites. With this we aim to emphasize a to date underestimated regulator that influences both proteases.


Assuntos
Proteína Morfogenética Óssea 1/química , Cloreto de Cálcio/química , Metaloendopeptidases/química , Proteína Morfogenética Óssea 1/metabolismo , Colágeno Tipo I/química , Colágeno Tipo I/metabolismo , Humanos , Metaloendopeptidases/metabolismo
19.
Sci Rep ; 7: 44053, 2017 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-28276471

RESUMO

Soluble Interleukin-6 receptor (sIL-6R) mediated trans-signaling is an important pro-inflammatory stimulus associated with pathological conditions, such as arthritis, neurodegeneration and inflammatory bowel disease. The sIL-6R is generated proteolytically from its membrane bound form and A Disintegrin And Metalloprotease (ADAM) 10 and 17 were shown to perform ectodomain shedding of the receptor in vitro and in vivo. However, under certain conditions not all sIL-6R could be assigned to ADAM10/17 activity. Here, we demonstrate that the IL-6R is a shedding substrate of soluble meprin α and membrane bound meprin ß, resulting in bioactive sIL-6R that is capable of inducing IL-6 trans-signaling. We determined cleavage within the N-terminal part of the IL-6R stalk region, distinct from the cleavage site reported for ADAM10/17. Interestingly, meprin ß can be shed from the cell surface by ADAM10/17 and the observation that soluble meprin ß is not capable of shedding the IL-6R suggests a regulatory mechanism towards trans-signaling. Additionally, we observed a significant negative correlation of meprin ß expression and IL-6R levels on human granulocytes, providing evidence for in vivo function of this proteolytic interaction.


Assuntos
Metaloendopeptidases/metabolismo , Receptores de Interleucina-6/metabolismo , Transdução de Sinais , Proteína ADAM10/genética , Proteína ADAM10/metabolismo , Proteína ADAM17/genética , Proteína ADAM17/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Secretases da Proteína Precursora do Amiloide/metabolismo , Células HEK293 , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Metaloendopeptidases/genética , Receptores de Interleucina-6/genética , Solubilidade
20.
PLoS Biol ; 15(1): e2000080, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-28060820

RESUMO

Signaling of the cytokine interleukin-6 (IL-6) via its soluble IL-6 receptor (sIL-6R) is responsible for the proinflammatory properties of IL-6 and constitutes an attractive therapeutic target, but how the sIL-6R is generated in vivo remains largely unclear. Here, we use liquid chromatography-mass spectrometry to identify an sIL-6R form in human serum that originates from proteolytic cleavage, map its cleavage site between Pro-355 and Val-356, and determine the occupancy of all O- and N-glycosylation sites of the human sIL-6R. The metalloprotease a disintegrin and metalloproteinase 17 (ADAM17) uses this cleavage site in vitro, and mutation of Val-356 is sufficient to completely abrogate IL-6R proteolysis. N- and O-glycosylation were dispensable for signaling of the IL-6R, but proteolysis was orchestrated by an N- and O-glycosylated sequon near the cleavage site and an N-glycan exosite in domain D1. Proteolysis of an IL-6R completely devoid of glycans is significantly impaired. Thus, glycosylation is an important regulator for sIL-6R generation.


Assuntos
Proteólise , Receptores de Interleucina-6/metabolismo , Proteína ADAM10/metabolismo , Proteína ADAM17/metabolismo , Processamento Alternativo/genética , Sequência de Aminoácidos , Secretases da Proteína Precursora do Amiloide/metabolismo , Linhagem Celular , Membrana Celular/metabolismo , Glicosilação , Humanos , Espaço Intracelular/metabolismo , Espectrometria de Massas , Proteínas de Membrana/metabolismo , Mutação/genética , Polissacarídeos/metabolismo , Prolina/metabolismo , Domínios Proteicos , Transporte Proteico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Interleucina-6/sangue , Receptores de Interleucina-6/química , Receptores de Interleucina-6/genética , Transdução de Sinais , Solubilidade , Valina/metabolismo
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