Granzyme B-induced and Caspase 3-dependent Cleavage of Gelsolin by Mouse Cytotoxic T Cells Modifies Cytoskeleton Dynamics

Martin, Praxedis; Pardo, Julián; Schill, Natalie; Jöckel, Lars; Berg, Matthias; Froelich, Christopher J.; Wallich, Reinhard; Simon, Markus M.

doi:10.1074/jbc.m109.056028

Cited by 17 publications

(14 citation statements)

References 54 publications

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“…Similar results were obtained using a second intracellular substrate of both caspase-3 and gzmB, that is gelsolin [24,25], which can be secreted and also be found in the plasma [26]. Plasma gelsolin from serum of C57BL/6 mice and recombinant gelsolin were cleaved by recombinant caspase-3, a proteolytic process specifically inhibited by caspase-3-specific inhibitor (Supporting Information Fig.…”

Section: Mc-released Active Caspase-3 Is Able To Cleave Native Il-33supporting

confidence: 75%

Secretory lysosomes of mouse mast cells store and exocytose active caspase‐3 in a strictly granzyme B dependent manner

et al. 2013

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In this study, we report that cytoplasmic granules from in vivo and in vitro derived mouse mast cells (MCs) contain active granzyme B (gzmB) and caspase-3, which is consistent with recent findings. Studying WT and gzmB-deficient mice, we observed that BM-derived MCs (BMMCs) from both strains contain cytosolic pro-caspase-3, but only WT BMMCs expressed active caspase-3 limited to their secretory lysosomes. Confocal microscopy revealed colocalization of active caspase-3 and gzmB in these cytoplasmic granules. The combined data demonstrate that the generation and storage of active caspase-3 is gzmBdependent. The finding that BMMCs secrete caspase-3 and gzmB after Ag stimulation suggests that both proteases contribute to extracellular MC-mediated proteolytic events. Although the extracellular function of MC-derived caspase-3 remains unclear, we show that BMMC-secreted caspase-3 cleaves IL-33, a cytokine that contributes to the development of asthma and arthritis. We also show that an in vitro propagated cytolytic T-lymphocyte line constitutively expresses gzmB together with active caspase-3, suggesting a novel interaction of these proteases in the execution of multiple innate and adaptive immune responses.Keywords: Caspase-3 r Cytotoxic T lymphocytes r Effector function r Granzyme B r Inflammation Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionMast cells (MCs) are essential effector cells in allergic disorders and other IgE-associated immune reactions [1]. Nevertheless, subsequent studies have demonstrated a much broader role for MCs in host defense and inflammation, ranging from the initiation Correspondence: Dr. Michael Huber e-mail: mhuber@ukaachen.de of innate immune responses to regulation of adaptive immunity [1][2][3]. MCs are distributed throughout all vascularized tissues residing in close proximity to blood vessels, nerves, and smooth muscle cells [4]. MCs are most commonly found at anatomical sites exposed to the environment, including the skin, airways, and the gastrointestinal tract [4]. * These authors contributed equally to this work. Eur. J. Immunol. 2013. 43: 3209-3218 MCs express numerous receptor classes, including . This cadre of sensors enables MCs to respond to multiple endogenous and exogenous stimuli releasing multiple biological mediators. Three main classes are distinguishable: (i) preformed mediators stored in secretory granules, that is proteoglycans and proteases such as tryptase, chymase, granzyme B (gzmB), and caspase-3 [5][6][7]; (ii) de novo generated lipid mediators, that is leukotrienes and prostaglandins; and (iii) a variety of cytokines, chemokines, and growth factors [4]. Cysteinedependent aspartate-directed proteases, namely caspases, initiate and execute the evolutionary conserved program of apoptosis [8]. Caspases are synthesized as zymogens with an N-terminal prodomain followed by sequences encoding a larger and then a smaller subunit [8]. Caspase-3 (apopain, CPP32), one of the so-called effecto...

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Section: Mc-released Active Caspase-3 Is Able To Cleave Native Il-33supporting

confidence: 75%

Secretory lysosomes of mouse mast cells store and exocytose active caspase‐3 in a strictly granzyme B dependent manner

et al. 2013

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“…Ϫ/Ϫ T cells are able to deliver granzyme(s), a recently described in vitro protocol was used (26,28). Here, gp33 pulsed or untreated MEF cells were incubated for 2 h in the presence or absence of recombinant gzmB plus LCMV-specific CD8 T cells from IL-1R Ϫ/Ϫ or perfxgzmAxB Ϫ/Ϫ mice.…”

Section: Perforin-expressing Lcmv-specific T Cells From Il-1r-deficiementioning

confidence: 99%

“…Exocytosis was analyzed as described previously (26,28). Accordingly, gp33 pulsed or untreated MEFs were incubated with or without recombinant gzmB and LCMV-specific CD8 T cells from either IL-1R Ϫ/Ϫ or perfxgzmAxB Ϫ/Ϫ mice for 2 h and analyzed for PS externalization and PI uptake.…”

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confidence: 99%

Interleukin-1R Signaling Is Essential for Induction of Proapoptotic CD8 T Cells, Viral Clearance, and Pathology during Lymphocytic Choriomeningitis Virus Infection in Mice

Joeckel

Wallich

Metkar

et al. 2012

J Virol

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gThe T cell granule exocytosis pathway is essential to control hepatotropic lymphocytic choriomeningitis virus strain WE (LCMV-WE) but also contributes to the observed pathology in mice. Although effective antiviral T cell immunity and development of viral hepatitis are strictly dependent on perforin and granzymes, the molecular basis underlying induction of functionally competent virus-immune T cells, including participation of the innate immune system, is far from being resolved. We demonstrate here that LCMV-immune T cells of interleukin-1 receptor (IL-1R)-deficient mice readily express transcripts for perforin and granzymes but only translate perforin, resulting in the lack of proapoptotic potential in vitro. LCMV is not cleared in IL-1R-deficient mice, and yet the infected mice develop neither splenomegaly nor hepatitis. These results demonstrate that IL-1R signaling is central to the induction of proapoptotic CD8 T cell immunity, including viral clearance and associated tissue injuries in LCMV infection.

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“…11 Nonetheless, a variety of studies have shown that hGrzB can also cleave other intracellular proteins with variable efficiency, and it is likely that some (or perhaps many) may have physiological relevance, including filamin, gelsolin and ROCKII. [12][13][14] Many viruses express Bcl-2-like proteins, and we have shown that the EBV orthologue BHRF1 is a potent blocker of hGrzB-mediated cell death. Despite this, intact cytotoxic lymphocytes (CTLs) (as distinct from purified recombinant reagents) are able to circumvent a Bcl-2-like block, 8 indicating that alternative pathways to cell death must exist.…”

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A functional genomics screen identifies PCAF and ADA3 as regulators of human granzyme B-mediated apoptosis and Bid cleavage

et al. 2014

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The human lymphocyte toxins granzyme B (hGrzB) and perforin cooperatively induce apoptosis of virus-infected or transformed cells: perforin pores enable entry of the serine protease hGrzB into the cytosol, where it processes Bid to selectively activate the intrinsic apoptosis pathway. Truncated Bid (tBid) induces Bax/Bak-dependent mitochondrial outer membrane permeability and the release of cytochrome c and Smac/Diablo. To identify cellular proteins that regulate perforin/hGrzB-mediated Bid cleavage and subsequent apoptosis, we performed a gene-knockdown (KD) screen using a lentiviral pool of short hairpin RNAs embedded within a miR30 backbone (shRNAmiR). We transduced HeLa cells with a lentiviral pool expressing shRNAmiRs that target 1213 genes known to be involved in cell death signaling and selected cells with acquired resistance to perforin/hGrzB-mediated apoptosis. Twenty-two shRNAmiRs were identified in the positive-selection screen including two, PCAF and ADA3, whose gene products are known to reside in the same epigenetic regulatory complexes. Small interfering (si)RNA-mediated gene-KD of PCAF or ADA3 also conferred resistance to perforin/hGrzB-mediated apoptosis providing independent validation of the screen results. Mechanistically, PCAF and ADA3 exerted their pro-apoptotic effect upstream of mitochondrial membrane permeabilization, as indicated by reduced cytochrome c release in PCAF-KD cells exposed to perforin/hGrzB. While overall levels of Bid were unaltered, perforin/hGrzB-mediated cleavage of Bid was reduced in PCAF-KD or ADA3-KD cells. We discovered that PCAF-KD or ADA3-KD resulted in reduced expression of PACS2, a protein implicated in Bid trafficking to mitochondria and importantly, targeted PACS2-KD phenocopied the effect of PCAF-KD or ADA3-KD. We conclude that PCAF and ADA3 regulate Bid processing via PACS2, to modulate the mitochondrial cell death pathway in response to hGrzB.

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Granzyme B-induced and Caspase 3-dependent Cleavage of Gelsolin by Mouse Cytotoxic T Cells Modifies Cytoskeleton Dynamics

Cited by 17 publications

References 54 publications

Secretory lysosomes of mouse mast cells store and exocytose active caspase‐3 in a strictly granzyme B dependent manner

Secretory lysosomes of mouse mast cells store and exocytose active caspase‐3 in a strictly granzyme B dependent manner

Interleukin-1R Signaling Is Essential for Induction of Proapoptotic CD8 T Cells, Viral Clearance, and Pathology during Lymphocytic Choriomeningitis Virus Infection in Mice

A functional genomics screen identifies PCAF and ADA3 as regulators of human granzyme B-mediated apoptosis and Bid cleavage

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