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A mutation in the kringle domain of human factor XII that causes autoinflammation, disturbs zymogen quiescence, and accelerates activation.

ABSTRACT: Coagulation factor XII (FXII) drives production of the inflammatory peptide bradykinin. Pathological mutations in the F12 gene, which encodes FXII, provoke acute tissue swelling in hereditary angioedema (HAE). Interestingly, a recently identified F12 mutation, causing a W268R substitution, is not associated with HAE. Instead, FXII-W268R carriers experience cold-inducible urticarial rash, arthralgia, fever, and fatigue. Here, we aimed to investigate the molecular characteristics of the FXII-W268R variant. We expressed wild type FXII (FXII-WT), FXII-W268R, and FXII-T309R (which causes HAE), as well as other FXII variants in HEK293 freestyle cells. Using chromogenic substrate assays, immunoblotting, and ELISA, we analyzed expression media, cell lysates, and purified proteins for FXII activation. Recombinant FXII-W268R forms increased amounts of intracellular cleavage products that are also present in expression medium and display enzymatic activity. The active site-incapacitated variant FXII-W268R/S544A reveals that intracellular fragmentation is largely dependent on autoactivation. Purified FXII-W268R is highly sensitive to activation by plasma kallikrein and plasmin, compared with FXII-WT or FXII-T309R. Furthermore, binding studies indicated that the FXII-W268R variant leads to the exposure of a plasminogen-binding site that is cryptic in FXII-WT. In plasma, recombinant FXII-W268R spontaneously triggers high-molecular-weight kininogen cleavage. Our findings suggest that the W268R substitution influences FXII protein conformation and exposure of the activation loop, which is concealed in FXII-WT. This results in intracellular autoactivation and constitutive low-grade secretion of activated FXII. These findings help to explain the chronically increased contact activation in carriers of the FXII-W268R variant.

SUBMITTER: Hofman ZLM

PROVIDER: S-EPMC6956518 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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A mutation in the kringle domain of human factor XII that causes autoinflammation, disturbs zymogen quiescence, and accelerates activation.

Hofman Zonne L M ZLM Clark Chantal C CC Sanrattana Wariya W Nosairi Aziz A Parr Naomi M J NMJ Živkovic Minka M Krause Karoline K Mahnke Niklas A NA Scheffel Jörg J Hack C Erik CE Maurer Marcus M de Maat Steven S Maas Coen C

The Journal of biological chemistry 20191126 2

Coagulation factor XII (FXII) drives production of the inflammatory peptide bradykinin. Pathological mutations in the <i>F12</i> gene, which encodes FXII, provoke acute tissue swelling in hereditary angioedema (HAE). Interestingly, a recently identified <i>F12</i> mutation, causing a W268R substitution, is not associated with HAE. Instead, FXII-W268R carriers experience cold-inducible urticarial rash, arthralgia, fever, and fatigue. Here, we aimed to investigate the molecular characteristics of ...[more]

PMID: 31771982

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Project description:Essentials During contact system activation, factor XII is progressively cleaved by plasma kallikrein. We investigated the role of factor XII truncation in biochemical studies. Factor XII contains naturally occurring truncating cleavage sites for a variety of enzymes. Truncation of factor XII primes it for activation in solution through exposure of R353. SUMMARY: Background The contact activation system and innate immune system are interlinked in inflammatory pathology. Plasma kallikrein (PKa) is held responsible for the stepwise processing of factor XII (FXII). A first cleavage activates FXII (into FXIIa); subsequent cleavages truncate it. This truncation eliminates its surface-binding domains, which negatively regulates surface-dependent coagulation. Objectives To investigate the influence of FXII truncation on its activation and downstream kallikrein-kinin system activation. Methods We study activation of recombinant FXII variants by chromogenic assays, by FXIIa ELISA and western blotting. Results We demonstrate that FXII truncation primes it for activation by PKa in solution. We demonstrate this phenomenon in three settings. (i) Truncation at a naturally occurring PKa-sensitive cleavage site, R334, accelerates FXIIa formation in solution. A site-directed mutant FXII-R334A displays ~50% reduced activity when exposed to PKa. (ii) A pathogenic mutation in FXII that causes hereditary angioedema, introduces an additional plasmin-sensitive cleavage site. Truncation at this site synergistically accelerates FXII activation in solution. (iii) We identify new, naturally occurring cleavage sites in FXII that have so far not been functionally linked to contact system activation. As examples, we show that non-activating truncation of FXII by neutrophil elastase and cathepsin K primes it for activation by PKa in solution. Conclusions FXII truncation, mediated by either pathogenic mutations or naturally occurring cleavage sites, primes FXII for activation in solution. We propose that the surface-binding domains of FXII shield its activating cleavage site, R353. This may help to explain how the contact system contributes to inflammatory pathology.

Dataset Information

A mutation in the kringle domain of human factor XII that causes autoinflammation, disturbs zymogen quiescence, and accelerates activation.

Publications

A mutation in the kringle domain of human factor XII that causes autoinflammation, disturbs zymogen quiescence, and accelerates activation.

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