The CAP1/Prss8 catalytic triad is not involved in PAR2 activation and protease nexin‐1 (PN‐1) inhibition

Crisante, Giovanna; Battista, Laura; Iwaszkiewicz, Justyna; Nesca, Valeria; Mérillat, Anne-Marie; Sergi, Chloé; Zoete, Vincent; Frateschi, Simona; Hummler, Edith

doi:10.1096/fj.14-253781

Cited by 14 publications

(21 citation statements)

References 69 publications

(103 reference statements)

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“…Likewise, catalytically inactive prostasin mutants can stimulate the activation of protease-activated receptor-2 in a reconstituted mammalian cell-based system (13). Strong support for a non-proteolytic function of prostasin in vivo has been gained from the observation that mis-expressed catalytically inactive prostasin induces severe skin pathology in transgenic mice (13,14), and in particular, by our recent demonstration that mice expressing only catalytically inactive endogenous prostasin, unlike prostasin null mice, display normal long-term survival (15).…”

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confidence: 99%

Distinct Developmental Functions of Prostasin (CAP1/PRSS8) Zymogen and Activated Prostasin

Friis

Madsen

Bugge

2016

Journal of Biological Chemistry

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The membrane-anchored serine prostasin (CAP1/PRSS8) is essential for barrier acquisition of the interfollicular epidermis and for normal hair follicle development. Consequently, prostasin null mice die shortly after birth. Prostasin is found in two forms in the epidermis: a one-chain zymogen and a two-chain proteolytically active form, generated by matriptase-dependent activation site cleavage. Here we used gene editing to generate mice expressing only activation site cleavage-resistant (zymogen-locked) endogenous prostasin. Interestingly, these mutant mice displayed normal interfollicular epidermal development and postnatal survival, but had defects in whisker and pelage hair formation. These findings identify two distinct in vivo functions of epidermal prostasin: a function in the interfollicular epidermis, not requiring activation site cleavage, that can be mediated by the zymogen-locked version of prostasin and a proteolysis-dependent function of activated prostasin in hair follicles, dependent on zymogen conversion by matriptase.Prostasin (also known as channel-activating protease-1, CAP1, and PRSS8) is a glycosylphosphatidylinositol-anchored trypsin-like serine protease that is widely expressed in epithelial tissues, including both the interfollicular and the follicular compartments of the epidermis. Loss-of-function genetic studies in mice have uncovered critical functions of prostasin in both the formation of epidermal barrier formation and the formation of whiskers and pelage hair (1, 2). Prostasin is synthesized as an inactive proform (zymogen) that is converted to a catalytically active protease by a single endoproteolytic cleavage in the conserved activation cleavage site (3-6). Prostasin zymogen conversion in the epidermis requires the membraneactivated serine protease matriptase (7-9). This observation, combined with the observation that both matriptase-deficient and prostasin-deficient mice display identical epidermal phenotypes, led to the formulation of the hypothesis that prostasin exerts its functions in this tissue as part of a matriptase-prostasin cell surface zymogen activation cascade (8). Several observations made during the last decade suggest, however, that prostasin may also execute biological functions independent of its own proteolytic activity. For example, in a reconstituted Xenopus oocyte system, prostasin can activate the epithelial sodium channel (ENaC) by inducing proteolytic cleavage of its ␥ subunit to release an inhibitory domain. However, this activation, which can be inhibited by the broad-spectrum serine protease inhibitor, aprotinin, can also be efficiently executed by mutant prostasin variants that lack the catalytic histidine-aspartate-serine triad (10 -12). Likewise, catalytically inactive prostasin mutants can stimulate the activation of protease-activated receptor-2 in a reconstituted mammalian cell-based system (13). Strong support for a non-proteolytic function of prostasin in vivo has been gained from the observation that mis-expressed catalytically inactive pros...

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mentioning

confidence: 99%

Distinct Developmental Functions of Prostasin (CAP1/PRSS8) Zymogen and Activated Prostasin

Friis

Madsen

Bugge

2016

Journal of Biological Chemistry

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“…doi:10.1371/journal.pone.0170944.g007 functions of prostasin zymogen identified via genetic manipulation in various model systems [39,[42][43][44][45].…”

Section: Discussionmentioning

confidence: 99%

“…In a study by Crisante et al [42] recombinant prostasin with mutations at all three amino acids of the active site triad was reported to form a stable complex with PN-1, a serpin-type inhibitor. Serpin inhibitors inactivate active proteases by the formation of acyl-enzyme intermediate involving the serine residue in the active site triad [51], and so PN-1 should not be able to form a stable complex with the mutant prostasin.…”

Section: Discussionmentioning

confidence: 99%

Selective Inhibition of Prostasin in Human Enterocytes by the Integral Membrane Kunitz-Type Serine Protease Inhibitor HAI-2

Yeo¹,

Shiao²,

Liu³

et al. 2017

Gastroenterology

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“…Although it remains elusive how FAPα exerts its functions in progression of these tumors it's evident, that a variety of different downstream pathways are involved [4]. There is increasing evidence of proteases being involved in protein complex formation [14][15][16]. We aimed at a holistic overview of the FAPα interactome and therefore applied the QUICK approach [18] using the CAF-derived cell line CT5.3.…”

Section: Overviewmentioning

confidence: 99%

“…At the same time, there is increasing evidence that proteases may exert their biological function in a non-enzymatic manner. Examples include involvement of matrix metalloproteases in cellular migration [14] and non-enzymatic functions of the cell surface serine protease prostasin [15,16]. Such observations make it essential to elucidate protease interactomes in order to better understand the molecular basis of their in vivo functions.…”

Section: Introductionmentioning

confidence: 99%

The stromal cell-surface protease fibroblast activation protein-α localizes to lipid rafts and is recruited to invadopodia

Knopf

Tholen

Koczorowska

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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Fibroblast activation protein alpha (FAPα) is a cell surface protease expressed by cancer-associated fibroblasts in the microenvironment of most solid tumors. As there is increasing evidence for proteases having non-catalytic functions, we determined the FAPα interactome in cancer-associated fibroblasts using the quantitative immunoprecipitation combined with knockdown (QUICK) method. Complex formation with adenosin deaminase, erlin-2, stomatin, prohibitin, Thy-1 membrane glycoprotein, and caveolin-1 was further validated by immunoblotting. Co-immunoprecipitation (co-IP) of the known stoichiometric FAPα binding partner dipeptidyl-peptidase IV (DPPIV) corroborated the proteomic strategy. Reverse co-IPs validated the FAPα interaction with caveolin-1, erlin-2, and stomatin while co-IP upon RNA-interference mediated knock-down of DPPIV excluded adenosin deaminase as a direct FAPα interaction partner. Many newly identified FAPα interaction partners localize to lipid rafts, including caveolin-1, a widely-used marker for lipid raft localization. We hypothesized that this indicates a recruitment of FAPα to lipid raft structures. In density gradient centrifugation, FAPα co-fractionates with caveolin-1. Immunofluorescence optical sectioning microscopy of FAPα and lipid raft markers further corroborates recruitment of FAPα to lipid rafts and invadopodia. FAPα is therefore an integral component of stromal lipid rafts in solid tumors. In essence, we provide one of the first interactome analyses of a cell surface protease and translate these results into novel biological aspects of a marker protein for cancer-associated fibroblasts.

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The CAP1/Prss8 catalytic triad is not involved in PAR2 activation and protease nexin‐1 (PN‐1) inhibition

Cited by 14 publications

References 69 publications

Distinct Developmental Functions of Prostasin (CAP1/PRSS8) Zymogen and Activated Prostasin

Distinct Developmental Functions of Prostasin (CAP1/PRSS8) Zymogen and Activated Prostasin

Selective Inhibition of Prostasin in Human Enterocytes by the Integral Membrane Kunitz-Type Serine Protease Inhibitor HAI-2

The stromal cell-surface protease fibroblast activation protein-α localizes to lipid rafts and is recruited to invadopodia

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