C5a receptor is cleaved by metalloproteases induced by sphingomyelinase D from Loxosceles spider venom

Berg, Carmen W. van den; Gonçalves-de-Andrade, Rute M.; Okamoto, Cinthya Kimori; Tambourgi, Denise V.

doi:10.1016/j.imbio.2012.01.005

Cited by 14 publications

(11 citation statements)

References 38 publications

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“…C5aR was detected as described (22) using mAb S5/1 or polyclonal anti-C5aR. Precision Plus All Blue standards (Bio-Rad, Hemel Hempstead, U.K.) were used to calculate the molecular weights of the bands.…”

Section: Sds-page and Western Blottingmentioning

confidence: 99%

“…Cell-surface C5aR expression was detected by flow cytometry as described (22) using mAb S5/1, and fluorescence was measured on an Accuri flow cytometer (BD Biosciences). Results are expressed as average of mean of arbitrary fluorescent intensity 6 SD of experiments carried out in triplicate.…”

Section: Flow Cytometrymentioning

confidence: 99%

“…After washing, 200 ml cells (5 3 10 6 /ml) was stimulated with 2.5 nM C5a or 250 nM C5aR agonist at 37˚C. Changes in intracellular calcium were measured as described (22,26).…”

Section: Calcium Flux Measurementsmentioning

confidence: 99%

“…NSPs have been shown to proteolytically reduce the expression of several cell-surface molecules involved in immunity, including CXCR1 (IL-8RA), CR1, CD16, CD43, and TNFRII, but their impact on C5aR expression has not been investigated. Proteolytic cleavage and inactivation of C5aR has been demonstrated by a variety of endogenous and exogenous proteases, including an endogenous metalloprotease, activated by the action of Loxosceles spider venom sphingomyelinase D, a serine protease from Porphyromonas gingivalis, and a metalloprotease from venom of the spider Plectreurys tristes (22)(23)(24). Direct proteolysis and inactivation of C5aR by an endogenous protease has not been demonstrated so far; however, a reduced chemotactic response to C5a after NE and CG treatment has been reported (25).…”

mentioning

confidence: 99%

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Mechanism of Neutrophil Dysfunction: Neutrophil Serine Proteases Cleave and Inactivate the C5a Receptor

Berg

Tambourgi

Clark

et al. 2014

The Journal of Immunology

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Neutrophil dysfunction, resulting in inefficient bacterial clearance, is a feature of several serious medical conditions, including cystic fibrosis (CF) and sepsis. Poorly controlled neutrophil serine protease (NSP) activity and complement activation have been implicated in this phenomenon. The capacity for excess NSP secretion and complement activation to influence the expression and function of the important neutrophil-activating receptor C5aR was investigated. Purified NSPs cathepsin G (CG), neutrophil elastase (NE), and proteinase 3 cleaved C5aR to a 26- to 27-kDa membrane-bound fragment, thereby inactivating its C5a-induced signaling ability. In a supernatant transfer assay, NSPs released from neutrophils in response to C5a induced the cleavage of the C5aR on unstimulated cells. Stimulation of myeolomonocytic U937 cells and purified neutrophils with C5a resulted in downregulation of the C5aR on these cells, which, in the case of U937 cells, was largely caused by NSP-mediated cleavage of C5aR, but in the case of neutrophils, intracellular degradation was likely the main mediator in addition to a small role for NSPs. CG and NE in bronchoalveolar lavage fluid from CF patients both contributed to C5aR cleavage. We propose two converging models for C5a- and NSP-mediated neutrophil dysfunction whereby C5aR cleavage is induced by NSPs, secreted in response to: 1) excess C5a generation or other stimuli; or 2) necrosis. The consequent impairment of C5aR activity contributes to suboptimal local neutrophil priming and bacterial clearance. NSP inhibitors with specificity for both CG and NE may aid the treatment of pathologies associated with neutrophil dysfunction including sepsis and CF.

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Section: Sds-page and Western Blottingmentioning

confidence: 99%

Section: Flow Cytometrymentioning

confidence: 99%

“…After washing, 200 ml cells (5 3 10 6 /ml) was stimulated with 2.5 nM C5a or 250 nM C5aR agonist at 37˚C. Changes in intracellular calcium were measured as described (22,26).…”

Section: Calcium Flux Measurementsmentioning

confidence: 99%

mentioning

confidence: 99%

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Mechanism of Neutrophil Dysfunction: Neutrophil Serine Proteases Cleave and Inactivate the C5a Receptor

Berg

Tambourgi

Clark

et al. 2014

The Journal of Immunology

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“…/ml) were stimulated with C5a, C5, or C5 preincubated with HNE (2 h at 37˚C as described above). Changes in intracellular calcium were measured as described previously (30,31). To confirm that the observed signal was C5a induced, a C5aR1 antagonist peptide (H-Phe-Lys-Pro-D-Cha-Trp-D-Arg-OH) (100 nM diluted in Krebs/HEPES/BSA) was added to the cells 10 min before stimulation with C5-derived cleavage products.…”

Section: Enzyme Cleavage Assaysmentioning

confidence: 99%

Functional analysis of a complement polymorphism (rs17611) associated with Rheumatoid Arthritis

Giles¹,

Morgan²,

Harris³

2013

Molecular Immunology

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Complement is implicated in the pathogenesis of rheumatoid arthritis (RA); elevated levels of complement activation products have been measured in plasma, synovial fluid, and synovial tissues of patients. Complement polymorphisms are associated with RA in genome-wide association studies. Coding-region polymorphisms may directly impact protein activity; indeed, we have shown that complement polymorphisms affecting a single amino acid change cause subtle changes in individual component function that in combination have dramatic effects on complement activity and disease risk. In this study, we explore the functional consequences of a single nucleotide polymorphism (SNP) (rs17611) encoding a V802I polymorphism in C5 and propose a mechanism for its link to RA pathology. Plasma levels of C5, C5a, and terminal complement complex were measured in healthy and RA donors and correlated to rs17611 polymorphic status. Impact of the SNP on C5 functionality was assessed. Plasma C5a levels were significantly increased and C5 levels significantly lower with higher copy number of the RA risk allele for rs17611, suggesting increased turnover of C5 V802. Functional assays using purified C5 variants revealed no significant differences in lytic activity, suggesting that increased C5 V802 turnover was not mediated by complement convertase enzymes. C5 is also cleaved in vivo by proteases; the C5 V802 variant was more sensitive to cleavage with elastase and the "C5a" generated was biologically active. We hypothesize that this SNP in C5 alters the rate at which elastase generates active C5a in rheumatoid joints, hence recruiting neutrophils to the site thus maintaining a state of inflammation in arthritic joints.

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Role of ErbB and IL-1 signaling pathways in the dermonecrotic lesion induced by Loxosceles sphingomyelinases D

Pinto,

Lopes,

Trufen

et al. 2023

Arch Toxicol

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C5a receptor is cleaved by metalloproteases induced by sphingomyelinase D from Loxosceles spider venom

Cited by 14 publications

References 38 publications

Mechanism of Neutrophil Dysfunction: Neutrophil Serine Proteases Cleave and Inactivate the C5a Receptor

Mechanism of Neutrophil Dysfunction: Neutrophil Serine Proteases Cleave and Inactivate the C5a Receptor

Functional analysis of a complement polymorphism (rs17611) associated with Rheumatoid Arthritis

Role of ErbB and IL-1 signaling pathways in the dermonecrotic lesion induced by Loxosceles sphingomyelinases D

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