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Chenyong Lin - One of the best experts on this subject based on the ideXlab platform.
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Targeted deletion of HAI-1 increases prostasin proteolysis but decreases Matriptase proteolysis in human keratinocytes
Human Cell, 2021Co-Authors: Robert J. Barndt, Jehng-kang Wang, Michael D. Johnson, Shih-ming Huang, Chenyong LinAbstract:Epidermal differentiation and barrier function require well-controlled Matriptase and prostasin proteolysis, in which the Kunitz-type serine protease inhibitor HAI-1 represents the primary enzymatic inhibitor for both proteases. HAI-1, however, also functions as a chaperone-like protein necessary for normal Matriptase synthesis and intracellular trafficking. Furthermore, other protease inhibitors, such as antithrombin and HAI-2, can also inhibit Matriptase and prostasin in solution or in keratinocytes. It remains unclear, therefore, whether aberrant increases in Matriptase and prostasin enzymatic activity would be the consequence of targeted deletion of HAI-1 and so subsequently contribute to the epidermal defects observed in HAI-1 knockout mice. The impact of HAI-1 deficiency on Matriptase and prostasin proteolysis was, here, investigated in HaCaT human keratinocytes. Our results show that HAI-1 deficiency causes an increase in prostasin proteolysis via increased protein expression and zymogen activation. It remains unclear, however, whether HAI-1 deficiency increases “net” prostasin enzymatic activity because all of the activated prostasin was detected in complexes with HAI-2, suggesting that prostasin enzymatic activity is still under tight control in HAI-1-deficient keratinocytes. Matriptase proteolysis is, however, unexpectedly suppressed by HAI-1 deficiency, as manifested by decreases in zymogen activation, shedding of active Matriptase, and Matriptase-dependent prostasin zymogen activation. This suppressed proteolysis results mainly from the reduced ability of HAI-1-deficient HaCaT cells to activate Matriptase and the rapid inhibition of nascent active Matriptase by HAI-2 and other yet-to-be-identified protease inhibitors. Our study provides novel insights with opposite impacts by HAI-1 deficiency on Matriptase versus prostasin proteolysis in keratinocytes.
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Mild acidity likely accelerates the physiological Matriptase autoactivation process: a comparative study between spontaneous and acid-induced Matriptase zymogen activation.
Human cell, 2020Co-Authors: Bailing Jia, Chenyong Lin, Yi Lin Chiu, Jehng-kang Wang, Robert B. Barndt, Hamishi A Thompson, Mon-juan Lee, See-chi Lee, Hung-jen Tang, Michael D. JohnsonAbstract:The pathophysiological functions of Matriptase, a type 2 transmembrane serine protease, rely primarily on its enzymatic activity, which is under tight control through multiple mechanisms. Among those regulatory mechanisms, the control of zymogen activation is arguably the most important. Matriptase zymogen activation not only generates the mature active enzyme but also initiates suppressive mechanisms, such as rapid inhibition by HAI-1, and Matriptase shedding. These tightly coupled events allow the potent Matriptase tryptic activity to fulfill its biological functions at the same time as limiting undesired hazards. Matriptase is converted to the active enzyme via a process of autoactivation, in which the activational cleavage is thought to rely on the interactions of Matriptase zymogen molecules and other as yet identified proteins. Matriptase autoactivation can occur spontaneously and is rapidly followed by the formation and then shedding of Matriptase-HAI-1 complexes, resulting in the presence of relatively low levels of the complex on cells. Activation can also be induced by several non-protease factors, such as the exposure of cells to a mildly acidic buffer, which rapidly causes high-level Matriptase zymogen activation in almost all cell lines tested. In the current study, the structural requirements for this acid-induced zymogen activation are compared with those required for spontaneous activation through a systematic analysis of the impact of 18 different mutations in various structural domains and motifs on Matriptase zymogen activation. Our study reveals that both acid-induced Matriptase activation and spontaneous activation depend on the maintenance of the structural integrity of the serine protease domain, non-catalytic domains, and posttranslational modifications. The common requirements of both modes of activation suggest that acid-induced Matriptase activation may function as a physiological mechanism to induce pericellular proteolysis by accelerating Matriptase autoactivation.
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the intracellular seven amino acid motif eegevfl is required for Matriptase vesicle sorting and translocation to the basolateral plasma membrane
PLOS ONE, 2020Co-Authors: Chun-che Tseng, Chenyong Lin, Bailing Jia, Jehng-kang Wang, Robert B. Barndt, Hung-jen Tang, Yanghong Dai, Yu Hsin Chen, Michael D. JohnsonAbstract:Matriptase plays important roles in epithelial integrity and function, which depend on its sorting to the basolateral surface of cells, where Matriptase zymogen is converted to an active enzyme in order to act on its substrates. After activation, Matriptase undergoes HAI-1-mediated inhibition, internalization, transcytosis, and secretion from the apical surface into the lumen. Matriptase is a mosaic protein with several distinct protein domains and motifs, which are a reflection of Matriptase's complex cellular itinerary, life cycle, and the tight control of its enzymatic activity. While the molecular determinants for various Matriptase regulatory events have been identified, the motif(s) required for translocation of human Matriptase to the basolateral plasma membrane is unknown. The motif previously identified in rat Matriptase is not conserved between the rodent and the primate. We, here, revisit the question for human Matriptase through the use of a fusion protein containing a green fluorescent protein linked to the Matriptase N-terminal fragment ending at Gly-149. A conserved seven amino acid motif EEGEVFL, which is similar to the monoleucine C-terminal to an acidic cluster motif involved in the basolateral targeting for some growth factors, has been shown to be required for Matriptase translocation to the basolateral plasma membrane of polarized MDCK cells. Furthermore, time-lapse video microscopy showed that the motif appears to be required for entry into the correct transport vesicles, by which Matriptase can undergo rapid trafficking and translocate to the plasma membrane. Our study reveals that the EEGEVFL motif is necessary, but may not be sufficient, for Matriptase basolateral membrane targeting and serves as the basis for further research on its pathophysiological roles.
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Matriptase shedding is closely coupled with Matriptase zymogen activation and requires de novo proteolytic cleavage likely involving its own activity.
PloS one, 2017Co-Authors: Chun-che Tseng, Bailing Jia, Robert J. Barndt, Chien-yu Chen, I-chu Tseng, Jehng-kang Wang, Michael D. Johnson, Chenyong LinAbstract:The type 2 transmembrane serine protease Matriptase is involved in many pathophysiological processes probably via its enzymatic activity, which depends on the dynamic relationship between zymogen activation and protease inhibition. Matriptase shedding can prolong the life of enzymatically active Matriptase and increase accessibility to substrates. We show here that Matriptase shedding occurs via a de novo proteolytic cleavage at sites located between the SEA domain and the CUB domain. Point or combined mutations at the four positively charged amino acid residues in the region following the SEA domain allowed Arg-186 to be identified as the primary cleavage site responsible for Matriptase shedding. Kinetic studies further demonstrate that Matriptase shedding is temporally coupled with Matriptase zymogen activation. The onset of Matriptase shedding lags one minute behind Matriptase zymogen activation. Studies with active site triad Ser-805 point mutated Matriptase, which no longer undergoes zymogen activation or shedding, further suggests that Matriptase shedding depends on Matriptase zymogen activation, and that Matriptase proteolytic activity may be involved in its own shedding. Our studies uncover an autonomous mechanism coupling Matriptase zymogen activation, proteolytic activity, and shedding such that a proportion of newly generated active Matriptase escapes HAI-1-mediated rapid inhibition by shedding into the extracellular milieu.
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Matriptase in complex HAI-1 is a shed species.
2017Co-Authors: Chun-che Tseng, Bailing Jia, Chien-yu Chen, I-chu Tseng, Jehng-kang Wang, Michael D. Johnson, Robert Barndt, Chenyong LinAbstract:HaCaT human keratinocyte cells were induced to activate Matriptase by pH 6.0 buffer treatment. Matriptase-HAI-1 complex in the lysate was immunodepleted with HAI-1 mAb-Sepharose. The cells lysate (lanes 1, 3, and 5) and the immunodepleted fraction (lanes 2, 4, and 6) were analyzed by western blot under non-reducing and non-boiled conditions for Matriptase CTF species (lanes 1 and 2), HAI-1 species (lanes 3 and 4), and Matriptase NTF (lanes 5 and 6). MTP stands for Matriptase, CTF for C terminal fragment, and NTF for N terminal fragment. The data presented are representative of at least 2 independent experiments.
Thomas H Bugge - One of the best experts on this subject based on the ideXlab platform.
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Matriptase zymogen supports epithelial development, homeostasis and regeneration.
BMC biology, 2017Co-Authors: Stine Friis, Katiuchia Uzzun Sales, Daniel Tadeo, Sylvain M. Le-gall, Henrik Jessen Jürgensen, Eric Camerer, Thomas H BuggeAbstract:Matriptase is a membrane serine protease essential for epithelial development, homeostasis, and regeneration, as well as a central orchestrator of pathogenic pericellular signaling in the context of inflammatory and proliferative diseases. Matriptase is an unusual protease in that its zymogen displays measurable enzymatic activity. Here, we used gain and loss of function genetics to investigate the possible biological functions of zymogen Matriptase. Unexpectedly, transgenic mice mis-expressing a zymogen-locked version of Matriptase in the epidermis displayed pathologies previously reported for transgenic mice mis-expressing wildtype epidermal Matriptase. Equally surprising, mice engineered to express only zymogen-locked endogenous Matriptase, unlike Matriptase null mice, were viable, developed epithelial barrier function, and regenerated the injured epithelium. Compatible with these observations, wildtype and zymogen-locked Matriptase were equipotent activators of PAR-2 inflammatory signaling. The study demonstrates that the Matriptase zymogen is biologically active and is capable of executing developmental and homeostatic functions of the protease.
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Matriptase zymogen supports epithelial development, homeostasis and regeneration
BMC Biology, 2017Co-Authors: Stine Friis, Katiuchia Uzzun Sales, Daniel Tadeo, Sylvain M. Le-gall, Henrik Jessen Jürgensen, Eric Camerer, Thomas H BuggeAbstract:Background Matriptase is a membrane serine protease essential for epithelial development, homeostasis, and regeneration, as well as a central orchestrator of pathogenic pericellular signaling in the context of inflammatory and proliferative diseases. Matriptase is an unusual protease in that its zymogen displays measurable enzymatic activity. Results Here, we used gain and loss of function genetics to investigate the possible biological functions of zymogen Matriptase. Unexpectedly, transgenic mice mis-expressing a zymogen-locked version of Matriptase in the epidermis displayed pathologies previously reported for transgenic mice mis-expressing wildtype epidermal Matriptase. Equally surprising, mice engineered to express only zymogen-locked endogenous Matriptase, unlike Matriptase null mice, were viable, developed epithelial barrier function, and regenerated the injured epithelium. Compatible with these observations, wildtype and zymogen-locked Matriptase were equipotent activators of PAR-2 inflammatory signaling. Conclusion The study demonstrates that the Matriptase zymogen is biologically active and is capable of executing developmental and homeostatic functions of the protease.
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HAI-2 stabilizes, inhibits and regulates SEA-cleavage-dependent secretory transport of Matriptase.
Traffic (Copenhagen Denmark), 2017Co-Authors: Annika W. Nonboe, Hiroaki Kataoka, Stine Friis, Signe Skovbjerg, Oliver Krigslund, Christoffer Soendergaard, Martin N. Andersen, Vincent Ellis, Makiko Kawaguchi, Thomas H BuggeAbstract:It has recently been shown that HAI-2 is able to suppress carcinogenesis induced by overexpression of Matriptase, as well as cause regression of individual established tumors in a mouse model system. However, the role of HAI-2 is poorly understood. In the present study we describe three mutations in the binding loop of the HAI-2 Kunitz domain 1 (K42N, C47F, and R48L) that cause a delay in the SEA domain cleavage of Matriptase, leading to accumulation of non-SEA domain cleaved Matriptase in the ER. We suggest that, like other known SEA domains, the Matriptase SEA domain auto-cleaves and reflects that correct oligomerization, maturation, and/or folding has been obtained. Our results suggest that the HAI-2 Kunitz domain 1 mutants influence the flux of Matriptase to the plasma membrane by affecting the oligomerization, maturation, and/or folding of Matriptase, and as a result the SEA domain cleavage of Matriptase. Two of the HAI-2 Kunitz domain 1 mutants investigated (C47F, R48L, C47F/R48L) also displayed a reduced ability to proteolytically silence Matriptase. Hence, HAI-2 separately stabilizes Matriptase, regulates the secretory transport, possibly via maturation/oligomerization, and inhibits the proteolytic activity of Matriptase in the ER, and possible throughout the secretory pathway.
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Matriptase promotes inflammatory cell accumulation and progression of established epidermal tumors.
Oncogene, 2014Co-Authors: Katiuchia Uzzun Sales, Stine Friis, Loreto Abusleme, Niki M. Moutsopoulos, Thomas H BuggeAbstract:Deregulation of Matriptase is a consistent feature of human epithelial cancers and correlates with poor disease outcome. We have previously shown that Matriptase promotes multi-stage squamous cell carcinogenesis in transgenic mice through dual activation of pro-hepatocyte growth factor-cMet-Akt-mTor proliferation/survival signaling and PAR-2-Gαi-NFκB inflammatory signaling. Matriptase was congenitally and constitutively deregulated in our prior studies, and therefore it was unclear if aberrant Matriptase signaling supports only initiation of tumor formation or if it is also critical for the progression of established tumors. To determine this, we here have generated triple-transgenic mice with constitutive deregulation of Matriptase and simultaneous inducible expression of the cognate Matriptase inhibitor, hepatocyte growth factor inhibitor (HAI)-2. As expected, constitutive expression of HAI-2 suppressed the formation of Matriptase-dependent tumors in 7,12-Dimethylbenz(a)anthracene (DMBA)-treated mouse skin. Interestingly, however, the induction of HAI-2 expression in already established tumors markedly impaired malignant progression and caused regression of individual tumors. Tumor regression correlated with reduced accumulation of tumor-associated inflammatory cells, likely caused by diminished expression of pro-tumorigenic inflammatory cytokines. The data suggest that Matriptase-dependent signaling may be a therapeutic target for both squamous cell carcinoma chemoprevention and for the treatment of established tumors.
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the protease inhibitor hai 2 but not hai 1 regulates Matriptase activation and shedding through prostasin
Journal of Biological Chemistry, 2014Co-Authors: Stine Friis, Hiroaki Kataoka, Katiuchia Uzzun Sales, Lotte K Vogel, Jeffrey Schafer, Thomas H BuggeAbstract:The membrane-anchored serine proteases, Matriptase and prostasin, and the membrane-anchored serine protease inhibitors, hepatocyte growth factor activator inhibitor (HAI)-1 and HAI-2, are critical effectors of epithelial development and postnatal epithelial homeostasis. Matriptase and prostasin form a reciprocal zymogen activation complex that results in the formation of active Matriptase and prostasin that are targets for inhibition by HAI-1 and HAI-2. Conflicting data, however, have accumulated as to the existence of auxiliary functions for both HAI-1 and HAI-2 in regulating the intracellular trafficking and activation of Matriptase. In this study, we, therefore, used genetically engineered mice to determine the effect of ablation of endogenous HAI-1 and endogenous HAI-2 on endogenous Matriptase expression, subcellular localization, and activation in polarized intestinal epithelial cells. Whereas ablation of HAI-1 did not affect Matriptase in epithelial cells of the small or large intestine, ablation of HAI-2 resulted in the loss of Matriptase from both tissues. Gene silencing studies in intestinal Caco-2 cell monolayers revealed that this loss of cell-associated Matriptase was mechanistically linked to accelerated activation and shedding of the protease caused by loss of prostasin regulation by HAI-2. Taken together, these data indicate that HAI-1 regulates the activity of activated Matriptase, whereas HAI-2 has an essential role in regulating prostasin-dependent Matriptase zymogen activation.
Michael D. Johnson - One of the best experts on this subject based on the ideXlab platform.
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Targeted deletion of HAI-1 increases prostasin proteolysis but decreases Matriptase proteolysis in human keratinocytes
Human Cell, 2021Co-Authors: Robert J. Barndt, Jehng-kang Wang, Michael D. Johnson, Shih-ming Huang, Chenyong LinAbstract:Epidermal differentiation and barrier function require well-controlled Matriptase and prostasin proteolysis, in which the Kunitz-type serine protease inhibitor HAI-1 represents the primary enzymatic inhibitor for both proteases. HAI-1, however, also functions as a chaperone-like protein necessary for normal Matriptase synthesis and intracellular trafficking. Furthermore, other protease inhibitors, such as antithrombin and HAI-2, can also inhibit Matriptase and prostasin in solution or in keratinocytes. It remains unclear, therefore, whether aberrant increases in Matriptase and prostasin enzymatic activity would be the consequence of targeted deletion of HAI-1 and so subsequently contribute to the epidermal defects observed in HAI-1 knockout mice. The impact of HAI-1 deficiency on Matriptase and prostasin proteolysis was, here, investigated in HaCaT human keratinocytes. Our results show that HAI-1 deficiency causes an increase in prostasin proteolysis via increased protein expression and zymogen activation. It remains unclear, however, whether HAI-1 deficiency increases “net” prostasin enzymatic activity because all of the activated prostasin was detected in complexes with HAI-2, suggesting that prostasin enzymatic activity is still under tight control in HAI-1-deficient keratinocytes. Matriptase proteolysis is, however, unexpectedly suppressed by HAI-1 deficiency, as manifested by decreases in zymogen activation, shedding of active Matriptase, and Matriptase-dependent prostasin zymogen activation. This suppressed proteolysis results mainly from the reduced ability of HAI-1-deficient HaCaT cells to activate Matriptase and the rapid inhibition of nascent active Matriptase by HAI-2 and other yet-to-be-identified protease inhibitors. Our study provides novel insights with opposite impacts by HAI-1 deficiency on Matriptase versus prostasin proteolysis in keratinocytes.
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Mild acidity likely accelerates the physiological Matriptase autoactivation process: a comparative study between spontaneous and acid-induced Matriptase zymogen activation.
Human cell, 2020Co-Authors: Bailing Jia, Chenyong Lin, Yi Lin Chiu, Jehng-kang Wang, Robert B. Barndt, Hamishi A Thompson, Mon-juan Lee, See-chi Lee, Hung-jen Tang, Michael D. JohnsonAbstract:The pathophysiological functions of Matriptase, a type 2 transmembrane serine protease, rely primarily on its enzymatic activity, which is under tight control through multiple mechanisms. Among those regulatory mechanisms, the control of zymogen activation is arguably the most important. Matriptase zymogen activation not only generates the mature active enzyme but also initiates suppressive mechanisms, such as rapid inhibition by HAI-1, and Matriptase shedding. These tightly coupled events allow the potent Matriptase tryptic activity to fulfill its biological functions at the same time as limiting undesired hazards. Matriptase is converted to the active enzyme via a process of autoactivation, in which the activational cleavage is thought to rely on the interactions of Matriptase zymogen molecules and other as yet identified proteins. Matriptase autoactivation can occur spontaneously and is rapidly followed by the formation and then shedding of Matriptase-HAI-1 complexes, resulting in the presence of relatively low levels of the complex on cells. Activation can also be induced by several non-protease factors, such as the exposure of cells to a mildly acidic buffer, which rapidly causes high-level Matriptase zymogen activation in almost all cell lines tested. In the current study, the structural requirements for this acid-induced zymogen activation are compared with those required for spontaneous activation through a systematic analysis of the impact of 18 different mutations in various structural domains and motifs on Matriptase zymogen activation. Our study reveals that both acid-induced Matriptase activation and spontaneous activation depend on the maintenance of the structural integrity of the serine protease domain, non-catalytic domains, and posttranslational modifications. The common requirements of both modes of activation suggest that acid-induced Matriptase activation may function as a physiological mechanism to induce pericellular proteolysis by accelerating Matriptase autoactivation.
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The spatiotemporal control of human Matriptase action on its physiological substrates: a case against a direct role for Matriptase proteolytic activity in profilaggrin processing and desquamation
Human Cell, 2020Co-Authors: Jehng-kang Wang, Michael D. JohnsonAbstract:Studies of human genetic disorders and animal models indicate that Matriptase plays essential roles in proteolytic processes associated with profilaggrin processing and desquamation at late stages of epidermal differentiation. The tissue distribution profile and zymogen activation status in human skin, however, suggests that Matriptase physiological function in the skin more likely lies in the proliferating and differentiating keratinocytes in the basal and spinous layers. Marked acanthosis with expanded spinous layer and lack of significant changes in intensity and expression pattern for several terminal differentiation markers in the skin of ARIH patients support Matriptase’s role in earlier rather than the later stages of differentiation. In addition to the tissue distribution, differential subcellular localization further limits the ability of extracellular Matriptase proteolytic activity to access the cytosolic non-membrane-bound keratohyalin granules, in which profilaggrin processing occurs. The short lifespan of active Matriptase, which results from tightly controlled zymogen activation, rapid inhibition by HAI-1, and shedding from cell surface, indicates that active Matriptase likely performs physiological functions via limited proteolysis on its substrates, as needed, rather than via a continuous bulk process. We, here, review these spatiotemporal controls of Matriptase proteolytic activity at the biochemical, cellular, and tissue level. Based on this in-depth understanding of how Matriptase activity is regulated, we argue that there is no direct involvement of Matriptase proteolytic activity in profilaggrin processing and desquamation. The defects in epidermal terminal differentiation associated with Matriptase deficiency are likely secondary and are due to putative disruption at earlier stages of differentiation.
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the intracellular seven amino acid motif eegevfl is required for Matriptase vesicle sorting and translocation to the basolateral plasma membrane
PLOS ONE, 2020Co-Authors: Chun-che Tseng, Chenyong Lin, Bailing Jia, Jehng-kang Wang, Robert B. Barndt, Hung-jen Tang, Yanghong Dai, Yu Hsin Chen, Michael D. JohnsonAbstract:Matriptase plays important roles in epithelial integrity and function, which depend on its sorting to the basolateral surface of cells, where Matriptase zymogen is converted to an active enzyme in order to act on its substrates. After activation, Matriptase undergoes HAI-1-mediated inhibition, internalization, transcytosis, and secretion from the apical surface into the lumen. Matriptase is a mosaic protein with several distinct protein domains and motifs, which are a reflection of Matriptase's complex cellular itinerary, life cycle, and the tight control of its enzymatic activity. While the molecular determinants for various Matriptase regulatory events have been identified, the motif(s) required for translocation of human Matriptase to the basolateral plasma membrane is unknown. The motif previously identified in rat Matriptase is not conserved between the rodent and the primate. We, here, revisit the question for human Matriptase through the use of a fusion protein containing a green fluorescent protein linked to the Matriptase N-terminal fragment ending at Gly-149. A conserved seven amino acid motif EEGEVFL, which is similar to the monoleucine C-terminal to an acidic cluster motif involved in the basolateral targeting for some growth factors, has been shown to be required for Matriptase translocation to the basolateral plasma membrane of polarized MDCK cells. Furthermore, time-lapse video microscopy showed that the motif appears to be required for entry into the correct transport vesicles, by which Matriptase can undergo rapid trafficking and translocate to the plasma membrane. Our study reveals that the EEGEVFL motif is necessary, but may not be sufficient, for Matriptase basolateral membrane targeting and serves as the basis for further research on its pathophysiological roles.
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Matriptase shedding is closely coupled with Matriptase zymogen activation and requires de novo proteolytic cleavage likely involving its own activity.
PloS one, 2017Co-Authors: Chun-che Tseng, Bailing Jia, Robert J. Barndt, Chien-yu Chen, I-chu Tseng, Jehng-kang Wang, Michael D. Johnson, Chenyong LinAbstract:The type 2 transmembrane serine protease Matriptase is involved in many pathophysiological processes probably via its enzymatic activity, which depends on the dynamic relationship between zymogen activation and protease inhibition. Matriptase shedding can prolong the life of enzymatically active Matriptase and increase accessibility to substrates. We show here that Matriptase shedding occurs via a de novo proteolytic cleavage at sites located between the SEA domain and the CUB domain. Point or combined mutations at the four positively charged amino acid residues in the region following the SEA domain allowed Arg-186 to be identified as the primary cleavage site responsible for Matriptase shedding. Kinetic studies further demonstrate that Matriptase shedding is temporally coupled with Matriptase zymogen activation. The onset of Matriptase shedding lags one minute behind Matriptase zymogen activation. Studies with active site triad Ser-805 point mutated Matriptase, which no longer undergoes zymogen activation or shedding, further suggests that Matriptase shedding depends on Matriptase zymogen activation, and that Matriptase proteolytic activity may be involved in its own shedding. Our studies uncover an autonomous mechanism coupling Matriptase zymogen activation, proteolytic activity, and shedding such that a proportion of newly generated active Matriptase escapes HAI-1-mediated rapid inhibition by shedding into the extracellular milieu.
Karin List - One of the best experts on this subject based on the ideXlab platform.
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The Matriptase-prostasin proteolytic cascade in epithelial development and pathology
Cell and Tissue Research, 2013Co-Authors: Gregory S. Miller, Karin ListAbstract:The type II transmembrane serine protease Matriptase has an essential role in the integrity and function of multiple epithelial tissues. In the epidermis, Matriptase activates the glycosylphosphatidylinositol (GPI) anchored membrane serine protease prostasin to initiate a proteolytic cascade that is required for the development of the stratum corneum barrier function. Accordingly, mice deficient for Matriptase phenocopy mice deficient for epidermal prostasin and present with impaired corneocyte differentiation, imparied lipid matrix formation, loss of profilaggrin processing and loss of tight junction formation and function. Together, these defects lead to a compromised epidermal barrier and result in fatal dehydration during the neonatal period. Proteolytic activity of the Matriptase-prostasin cascade is regulated in the epidermis via inhibition by the Kunitz-type serine protease inhibitor hepatocyte growth factor activator inhibitor-1 (HAI-1). Importantly, targeted post-natal ablation of Matriptase in mice perturbs the function of multiple adult tissues, indicating an ongoing requirement for Matriptase proteolysis in the maintenance of diverse types of epithelia. Impaired Matriptase proteolytic activity has been linked to human Autosomal Recessive Icthyosis with Hypotrichosis (ARIH), whereas aberrant Matriptase activity has been implicated in Netherton’s Syndrome. This review will summarize information pertaining to the role of Matriptase in epithelial biology and will discuss recent advancements in our understanding of how Matriptase activity is regulated and the down-stream effectors of Matriptase proteolysis.
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membrane anchored serine protease Matriptase regulates epithelial barrier formation and permeability in the intestine
Proceedings of the National Academy of Sciences of the United States of America, 2010Co-Authors: Marguerite S. Buzza, Chenyong Lin, Karin List, Roman Szabo, Thomas H Bugge, Sarah Netzelarnett, Terez Sheadonohue, Aiping Zhao, Alessio Fasano, Toni M. AntalisAbstract:The intestinal epithelium serves as a major protective barrier between the mammalian host and the external environment. Here we show that the transmembrane serine protease Matriptase plays a pivotol role in the formation and integrity of the intestinal epithelial barrier. St14 hypomorphic mice, which have a 100-fold reduction in intestinal Matriptase mRNA levels, display a 35% reduction in intestinal transepithelial electrical resistance (TEER). Matriptase is expressed during intestinal epithelial differentiation and colocalizes with E-cadherin to apical junctional complexes (AJC) in differentiated polarized Caco-2 monolayers. Inhibition of Matriptase activity using a specific peptide inhibitor or by knockdown of Matriptase by siRNA disrupts the development of TEER in barrier-forming Caco-2 monolayers and increases paracellular permeability to macromolecular FITC-dextran. Loss of Matriptase was associated with enhanced expression and incorporation of the permeability-associated, “leaky” tight junction protein claudin-2 at intercellular junctions. Knockdown of claudin-2 enhanced the development of TEER in Matriptase-silenced Caco-2 monolayers, suggesting that the reduced barrier integrity was caused, at least in part, by an inability to regulate claudin-2 expression and incorporation into junctions. We find that Matriptase enhances the rate of claudin-2 protein turnover, and that this is mediated indirectly through an atypical PKCζ-dependent signaling pathway. These results support a key role for Matriptase in regulating intestinal epithelial barrier competence, and suggest an intriguing link between pericellular serine protease activity and tight junction assembly in polarized epithelia.
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potent inhibition and global co localization implicate the transmembrane kunitz type serine protease inhibitor hepatocyte growth factor activator inhibitor 2 in the regulation of epithelial Matriptase activity
Journal of Biological Chemistry, 2008Co-Authors: Roman Szabo, Karin List, Alfredo A Molinolo, John P Hobson, Thomas H BuggeAbstract:Hepatocyte growth factor activator inhibitors (HAI)-1 and -2 are recently identified and closely related Kunitz-type transmembrane serine protease inhibitors. Whereas HAI-1 is well established as an inhibitor of the serine proteases Matriptase and hepatocyte growth factor activator, the physiological targets of HAI-2 are unknown. Here we show that HAI-2 displays potent inhibitory activity toward Matriptase, forms SDS-stable complexes with the serine protease, and blocks Matriptase-dependent activation of its candidate physiological substrates proprostasin and cell surface-bound pro-urokinase plasminogen activator. To further explore the potential functional relationship between HAI-2 and Matriptase, we generated a transgenic mouse strain with a promoterless β-galactosidase marker gene inserted into the endogenous locus encoding HAI-2 protein and performed a global high resolution mapping of the expression of HAI-2, Matriptase, and HAI-1 proteins in all adult tissues. This analysis showed striking co-localization of HAI-2 with Matriptase and HAI-1 in epithelial cells of all major organ systems, thus strongly supporting a role of HAI-2 as a physiological regulator of Matriptase activity, possibly acting in a redundant or partially redundant manner with HAI-1. Unlike HAI-1 and Matriptase, however, HAI-2 expression was also detected in non-epithelial cells of brain and lymph nodes, suggesting that HAI-2 may also be involved in inhibition of serine proteases other than Matriptase.
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co localization of the channel activating protease prostasin cap1 prss8 with its candidate activator Matriptase
Journal of Cellular Physiology, 2007Co-Authors: Karin List, Alfredo A Molinolo, John P Hobson, Thomas H BuggeAbstract:Prostasin (CAP1/PRSS8) is a glycosylphosphatidylinositol-anchored membrane serine protease believed to be critical for the regulation of epithelial sodium channel (ENaC) activity. Prostasin is synthesized as an inactive zymogen that requires a site-specific endoproteolytic cleavage to be converted to an active protease. We have recently reported that the tumor-associated type II transmembrane serine protease, Matriptase is necessary and sufficient for prostasin activation in the epidermis. In this study, the interrelationship between the two membrane serine proteases was investigated further by using enzymatic gene trapping combined with immunohistochemistry to delineate the spatial expression of Matriptase and prostasin in mouse tissues. We utilized a knock-in mouse with a promoterless beta-galactosidase marker gene inserted into the Matriptase locus, as a unique tool for precise assessment of endogenous Matriptase expression. The spatial expression of Matriptase and prostasin in mouse tissues was delineated by combining in situ beta-galactosidase Matriptase staining with immunohistochemical detection of prostasin. We report that prostasin displays a near-ubiquitous co-localization with its candidate activator Matriptase in a variety of normal epithelial tissues. These include simple, stratified, and pseudo-stratified epithelium of the integumentary system, digestive tract, respiratory tract, and urogenital tract. However, Matriptase and prostasin expression segregates during epithelial multi-stage carcinogenesis to eventually become localized in separate compartments of the tumor. These data suggest that a Matriptase-prostasin zymogen activation cascade may be functionally operative in multiple epithelial tissues, but Matriptase promotes epithelial carcinogenesis independent of prostasin.
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Matriptase-dependent cell surface proteolysis in epithelial development and pathogenesis.
Frontiers in bioscience : a journal and virtual library, 2007Co-Authors: Thomas H Bugge, Karin List, Roman SzaboAbstract:Matriptase is an epithelial type II transmembrane serine protease with a complex modular structure and sophisticated activation mechanism. Reduced Matriptase activity in mice or humans is associated with incomplete terminal differentiation of epidermis, epidermal appendages, oral epithelium, and, likely, other epithelial structures. Preliminary evidence indicates that Matriptase is part of a serine protease zymogen activation cascade that regulates epithelial cell proliferation and fate. Matriptase activity must be tightly controlled in epithelial tissues by transcriptional and posttranslational mechanisms, as Matriptase dysregulation can cause embryonic demise as well as malignant transformation.
Roman Szabo - One of the best experts on this subject based on the ideXlab platform.
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reduced prostasin cap1 prss8 activity eliminates hai 1 and hai 2 deficiency associated developmental defects by preventing Matriptase activation
PLOS Genetics, 2012Co-Authors: Roman Szabo, Katiuchia Uzzun Sales, Stine Friis, Sine Godiksen, Karina K Hansen, Peter Kosa, Natalia A Shylo, Silvio J GutkindAbstract:Loss of either hepatocyte growth factor activator inhibitor (HAI)-1 or -2 is associated with embryonic lethality in mice, which can be rescued by the simultaneous inactivation of the membrane-anchored serine protease, Matriptase, thereby demonstrating that a Matriptase-dependent proteolytic pathway is a critical developmental target for both protease inhibitors. Here, we performed a genetic epistasis analysis to identify additional components of this pathway by generating mice with combined deficiency in either HAI-1 or HAI-2, along with genes encoding developmentally co-expressed candidate Matriptase targets, and screening for the rescue of embryonic development. Hypomorphic mutations in Prss8, encoding the GPI-anchored serine protease, prostasin (CAP1, PRSS8), restored placentation and normal development of HAI-1–deficient embryos and prevented early embryonic lethality, mid-gestation lethality due to placental labyrinth failure, and neural tube defects in HAI-2–deficient embryos. Inactivation of genes encoding c-Met, protease-activated receptor-2 (PAR-2), or the epithelial sodium channel (ENaC) alpha subunit all failed to rescue embryonic lethality, suggesting that deregulated Matriptase-prostasin activity causes developmental failure independent of aberrant c-Met and PAR-2 signaling or impaired epithelial sodium transport. Furthermore, phenotypic analysis of PAR-1 and Matriptase double-deficient embryos suggests that the protease may not be critical for focal proteolytic activation of PAR-2 during neural tube closure. Paradoxically, although Matriptase auto-activates and is a well-established upstream epidermal activator of prostasin, biochemical analysis of Matriptase- and prostasin-deficient placental tissues revealed a requirement of prostasin for conversion of the Matriptase zymogen to active Matriptase, whereas prostasin zymogen activation was Matriptase-independent.
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membrane anchored serine protease Matriptase regulates epithelial barrier formation and permeability in the intestine
Proceedings of the National Academy of Sciences of the United States of America, 2010Co-Authors: Marguerite S. Buzza, Chenyong Lin, Karin List, Roman Szabo, Thomas H Bugge, Sarah Netzelarnett, Terez Sheadonohue, Aiping Zhao, Alessio Fasano, Toni M. AntalisAbstract:The intestinal epithelium serves as a major protective barrier between the mammalian host and the external environment. Here we show that the transmembrane serine protease Matriptase plays a pivotol role in the formation and integrity of the intestinal epithelial barrier. St14 hypomorphic mice, which have a 100-fold reduction in intestinal Matriptase mRNA levels, display a 35% reduction in intestinal transepithelial electrical resistance (TEER). Matriptase is expressed during intestinal epithelial differentiation and colocalizes with E-cadherin to apical junctional complexes (AJC) in differentiated polarized Caco-2 monolayers. Inhibition of Matriptase activity using a specific peptide inhibitor or by knockdown of Matriptase by siRNA disrupts the development of TEER in barrier-forming Caco-2 monolayers and increases paracellular permeability to macromolecular FITC-dextran. Loss of Matriptase was associated with enhanced expression and incorporation of the permeability-associated, “leaky” tight junction protein claudin-2 at intercellular junctions. Knockdown of claudin-2 enhanced the development of TEER in Matriptase-silenced Caco-2 monolayers, suggesting that the reduced barrier integrity was caused, at least in part, by an inability to regulate claudin-2 expression and incorporation into junctions. We find that Matriptase enhances the rate of claudin-2 protein turnover, and that this is mediated indirectly through an atypical PKCζ-dependent signaling pathway. These results support a key role for Matriptase in regulating intestinal epithelial barrier competence, and suggest an intriguing link between pericellular serine protease activity and tight junction assembly in polarized epithelia.
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potent inhibition and global co localization implicate the transmembrane kunitz type serine protease inhibitor hepatocyte growth factor activator inhibitor 2 in the regulation of epithelial Matriptase activity
Journal of Biological Chemistry, 2008Co-Authors: Roman Szabo, Karin List, Alfredo A Molinolo, John P Hobson, Thomas H BuggeAbstract:Hepatocyte growth factor activator inhibitors (HAI)-1 and -2 are recently identified and closely related Kunitz-type transmembrane serine protease inhibitors. Whereas HAI-1 is well established as an inhibitor of the serine proteases Matriptase and hepatocyte growth factor activator, the physiological targets of HAI-2 are unknown. Here we show that HAI-2 displays potent inhibitory activity toward Matriptase, forms SDS-stable complexes with the serine protease, and blocks Matriptase-dependent activation of its candidate physiological substrates proprostasin and cell surface-bound pro-urokinase plasminogen activator. To further explore the potential functional relationship between HAI-2 and Matriptase, we generated a transgenic mouse strain with a promoterless β-galactosidase marker gene inserted into the endogenous locus encoding HAI-2 protein and performed a global high resolution mapping of the expression of HAI-2, Matriptase, and HAI-1 proteins in all adult tissues. This analysis showed striking co-localization of HAI-2 with Matriptase and HAI-1 in epithelial cells of all major organ systems, thus strongly supporting a role of HAI-2 as a physiological regulator of Matriptase activity, possibly acting in a redundant or partially redundant manner with HAI-1. Unlike HAI-1 and Matriptase, however, HAI-2 expression was also detected in non-epithelial cells of brain and lymph nodes, suggesting that HAI-2 may also be involved in inhibition of serine proteases other than Matriptase.
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Matriptase-dependent cell surface proteolysis in epithelial development and pathogenesis.
Frontiers in bioscience : a journal and virtual library, 2007Co-Authors: Thomas H Bugge, Karin List, Roman SzaboAbstract:Matriptase is an epithelial type II transmembrane serine protease with a complex modular structure and sophisticated activation mechanism. Reduced Matriptase activity in mice or humans is associated with incomplete terminal differentiation of epidermis, epidermal appendages, oral epithelium, and, likely, other epithelial structures. Preliminary evidence indicates that Matriptase is part of a serine protease zymogen activation cascade that regulates epithelial cell proliferation and fate. Matriptase activity must be tightly controlled in epithelial tissues by transcriptional and posttranslational mechanisms, as Matriptase dysregulation can cause embryonic demise as well as malignant transformation.
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delineation of Matriptase protein expression by enzymatic gene trapping suggests diverging roles in barrier function hair formation and squamous cell carcinogenesis
American Journal of Pathology, 2006Co-Authors: Karin List, Roman Szabo, Alfredo A Molinolo, Boye Schnack Nielsen, Thomas H BuggeAbstract:The membrane serine protease Matriptase is required for epidermal barrier function, hair formation, and thymocyte development in mice, and dysregulated Matriptase expression causes epidermal squamous cell carcinoma. To elucidate the specific functions of Matriptase in normal and aberrant epidermal differentiation, we used enzymatic gene trapping combined with immunohistochemical, ultrastructural, and barrier function assays to delineate the spatio-temporal expression and function of Matriptase in mouse keratinized tissue development, homeostasis, and malignant transformation. In the interfollicular epidermis, Matriptase expression was restricted to postmitotic transitional layer keratinocytes undergoing terminal differentiation. Matriptase was also expressed in keratinizing oral epithelium, where it was required for oral barrier function, and in thymic epithelium. In all three tissues, Matriptase colocalized with profilaggrin. In staged embryos, the onset of epidermal Matriptase expression coincided with that of profilaggrin expression and acquisition of the epidermal barrier. In marked contrast to stratifying keritinized epithelium, matripase expression commenced already in undifferentiated and rapidly proliferating profilaggrin-negative matrix cells and displayed hair growth cycle-dependent expression. Exposure of the epidermis to carcinogens led to the gradual appearance of Matriptase in a keratin-5-positive proliferative cell compartment during malignant progression. Combined with previous studies, these data suggest that Matriptase has diverging functions in the genesis of stratified keratinized epithelium, hair follicles, and squamous cell carcinoma.