Calcium-independent phospholipase A2 (iPLA2) has been suggested to play an important role in the activation of caspase-1 induced by lipopolysaccharides (LPS). Here, we used pharmacological and genetic approaches to study the role of iPLA2 in the activation of caspase-1. Bromoenol lactone (BEL), an inhibitor that was originally used to support a role for iPLA2 in the secretion of IL-1β, prevented caspase-1 activation induced by LPS and ATP as described, and also activation triggered by Salmonella infection and cytosolic flagellin, which rely on the Nlrc4 inflammasome. Analysis of BEL enantiomers showed that the S-BEL form was more effective than R-BEL in inhibiting the inflammasome, suggesting a role for iPLA2β. However, caspase-1 activation and IL-1β secretion and their inhibition by BEL were unimpaired in macrophages deficient in iPLA2β. BEL was originally identified as an inhibitor of serine proteases. Consistent with the latter, the serine proteases inhibitors TPCK, TLCK and AAF-cmk prevented the activation of the Nlrc4 and Nlrp3 inflammasomes while pan-cathepsin inhibitors were ineffective. These results indicate that iPLA2β is not critical for caspase-1 activation as currently proposed. Instead, the results suggest that serine protease(s) targeted by BEL may play a critical role in the activation of the inflammasome triggered by microbial stimuli.
© 2009 S. Karger AG, Basel
- Phospholipase A2
- Burns K, Martinon F, Tschopp J: New insights into the mechanism of IL-1beta maturation. Curr Opin Immunol 2003;15:26–30.
- Franchi L, Eigenbrod T, Munoz-Planillo R, Nunez G: The inflammasome: a caspase-1-activation platform that regulates immune responses and disease pathogenesis. Nat Immunol 2009;10:241–247.
- Franchi L, Park JH, Shaw MH, Marina-Garcia N, Chen G, Kim YG, Nunez G: Intracellular NOD-like receptors in innate immunity, infection and disease. Cell Microbiol 2008;10:1–8.
- Franchi L, McDonald C, Kanneganti TD, Amer A, Nunez G: Nucleotide-binding oligomerization domain-like receptors: intracellular pattern recognition molecules for pathogen detection and host defense. J Immunol 2006;177:3507–3513.
- Martinon F, Tschopp J: Inflammatory caspases and inflammasomes: master switches of inflammation. Cell Death Differ 2007;14:10–22.
- Franchi L, Warner N, Viani K, Nunez G: Function of NOD-like receptors in microbial recognition and host defense. Immunol Rev 2009;227:106–128.
- Schaloske RH, Dennis EA: The phospholipase A2 superfamily and its group numbering system. Biochim Biophys Acta 2006;1761:1246–1259.
- Qu Y, Franchi L, Nunez G, Dubyak GR: Nonclassical IL-1 beta secretion stimulated by P2X7 receptors is dependent on inflammasome activation and correlated with exosome release in murine macrophages. J Immunol 2007;179:1913–1925.
- Kahlenberg JM, Dubyak GR: Mechanisms of caspase-1 activation by P2X7 receptor-mediated K+ release. Am J Physiol Cell Physiol 2004;286:C1100–1108.
- Andrei C, Margiocco P, Poggi A, Lotti LV, Torrisi MR, Rubartelli A: Phospholipases C and A2 control lysosome-mediated IL-1 beta secretion: implications for inflammatory processes. Proc Natl Acad Sci USA 2004;101:9745–9750.
- Walev I, Klein J, Husmann M, Valeva A, Strauch S, Wirtz H, Weichel O, Bhakdi S: Potassium regulates IL-1 beta processing via calcium-independent phospholipase A2. J Immunol 2000;164:5120–5124.
- Ackermann EJ, Conde-Frieboes K, Dennis EA: Inhibition of macrophage Ca(2+)-independent phospholipase A2 by bromoenol lactone and trifluoromethyl ketones. J Biol Chem 1995;270:445–450.
- Song H, Ramanadham S, Bao S, Hsu FF, Turk J: A bromoenol lactone suicide substrate inactivates group via phospholipase A2 by generating a diffusible bromomethyl keto acid that alkylates cysteine thiols. Biochemistry 2006;45:1061–1073.
- Daniels SB, Cooney E, Sofia MJ, Chakravarty PK, Katzenellenbogen JA: Haloenol lactones: potent enzyme-activated irreversible inhibitors for alpha-chymotrypsin. J Biol Chem 1983;258:15046–15053.
- Bao S, Miller DJ, Ma Z, Wohltmann M, Eng G, Ramanadham S, Moley K, Turk J: Male mice that do not express group via phospholipase A2 produce spermatozoa with impaired motility and have greatly reduced fertility. J Biol Chem 2004;279:38194–38200.
- Mariathasan S, Weiss DS, Newton K, McBride J, O’Rourke K, Roose-Girma M, Lee WP, Weinrauch Y, Monack DM, Dixit VM: Cryopyrin activates the inflammasome in response to toxins and ATP. Nature 2006;440:228–232.
- Sutterwala FS, Ogura Y, Szczepanik M, Lara-Tejero M, Lichtenberger GS, Grant EP, Bertin J, Coyle AJ, Galan JE, Askenase PW, Flavell RA: Critical role for NALP3/CIAS1/Cryopyrin in innate and adaptive immunity through its regulation of caspase-1. Immunity 2006;24:317–327.
- Fuentes L, Perez R, Nieto ML, Balsinde J, Balboa MA: Bromoenol lactone promotes cell death by a mechanism involving phosphatidate phosphohydrolase-1 rather than calcium-independent phospholipase A2. J Biol Chem 2003;278:44683–44690.
- Franchi L, Amer A, Body-Malapel M, Kanneganti TD, Ozoren N, Jagirdar R, Inohara N, Vandenabeele P, Bertin J, Coyle A, Grant EP, Nunez G: Cytosolic flagellin requires Ipaf for activation of caspase-1 and interleukin 1beta in Salmonella-infected macrophages. Nat Immunol 2006;7:576–582.
- Miao EA, Alpuche-Aranda CM, Dors M, Clark AE, Bader MW, Miller SI, Aderem A: Cytoplasmic flagellin activates caspase-1 and secretion of interleukin 1beta via Ipaf. Nat Immunol 2006;7:569–575.
- Moran JM, Buller RM, McHowat J, Turk J, Wohltmann M, Gross RW, Corbett JA: Genetic and pharmacologic evidence that calcium-independent phospholipase A2beta regulates virus-induced inducible nitric-oxide synthase expression by macrophages. J Biol Chem 2005;280:28162–28168.
- Jenkins CM, Han X, Mancuso DJ, Gross RW: Identification of calcium-independent phospholipase A2 (iPLA2) beta, and not iPLA2gamma, as the mediator of arginine vasopressin-induced arachidonic acid release in A-10 smooth muscle cells: enantioselective mechanism-based discrimination of mammalian iPLA2S. J Biol Chem 2002;277:32807–32814.
- Bao S, Li Y, Lei X, Wohltmann M, Jin W, Bohrer A, Semenkovich CF, Ramanadham S, Tabas I, Turk J: Attenuated free cholesterol loading-induced apoptosis but preserved phospholipid composition of peritoneal macrophages from mice that do not express group via phospholipase A2. J Biol Chem 2007;282:27100–27114.
- Xie Z, Gong MC, Su W, Turk J, Guo Z: Group via phospholipase A2 (iPLA2beta) participates in angiotensin II-induced transcriptional up-regulation of regulator of G-protein signaling-2 in vascular smooth muscle cells. J Biol Chem 2007;282:25278–25289.
- Lamkanfi M, Kanneganti TD, Franchi L, Nunez G: Caspase-1 inflammasomes in infection and inflammation. J Leukoc Biol 2007;82:220–225.
- Nicholson DW: Caspase structure, proteolytic substrates, and function during apoptotic cell death. Cell Death Differ 1999;6:1028–1042.
- Dowds TA, Masumoto J, Zhu L, Inohara N, Nunez G: Cryopyrin-induced interleukin 1beta secretion in monocytic cells: enhanced activity of disease-associated mutants and requirement for ASC. J Biol Chem 2004;279:21924–21928.
- Martinon F, Burns K, Tschopp J: The inflammasome: a molecular platform triggering activation of inflammatory caspases and processing of proIL-beta. Mol Cell 2002;10:417–426.
- Hilbi H, Puro RJ, Zychlinsky A: Tripeptidyl peptidase II promotes maturation of caspase-1 in Shigella flexneri-induced macrophage apoptosis. Infect Immun 2000;68:5502–5508.
- Suzuki T, Franchi L, Toma C, Ashida H, Ogawa M, Yoshikawa Y, Mimuro H, Inohara N, Sasakawa C, Nunez G: Differential regulation of caspase-1 activation, pyroptosis, and autophagy via Ipaf and ASC in Shigella-infected macrophages. PLoS Pathog 2007;3:e111.
- Marina-Garcia N, Franchi L, Kim YG, Miller D, McDonald C, Boons GJ, Nunez G: Pannexin-1-mediated intracellular delivery of muramyl dipeptide induces caspase-1 activation via cryopyrin/NLRP3 independently of Nod2. J Immunol 2008;180:4050–4057.
- Kanneganti TD, Lamkanfi M, Kim YG, Chen G, Park JH, Franchi L, Vandenabeele P, Nunez G: Pannexin-1-mediated recognition of bacterial molecules activates the cryopyrin inflammasome independent of toll-like receptor signaling. Immunity 2007;26:433–443.
- Ozoren N, Masumoto J, Franchi L, Kanneganti TD, Body-Malapel M, Erturk I, Jagirdar R, Zhu L, Inohara N, Bertin J, Coyle A, Grant EP, Nunez G: Distinct roles of TLR2 and the adaptor ASC in IL-1beta/IL-18 secretion in response to Listeria monocytogenes. J Immunol 2006;176:4337–4342.
- Kanneganti TD, Ozoren N, Body-Malapel M, Amer A, Park JH, Franchi L, Whitfield J, Barchet W, Colonna M, Vandenabeele P, Bertin J, Coyle A, Grant EP, Akira S, Nunez G: Bacterial RNA and small antiviral compounds activate caspase-1 through cryopyrin/Nalp3. Nature 2006;440:233–236.
- Kanneganti TD, Body-Malapel M, Amer A, Park JH, Whitfield J, Franchi L, Taraporewala ZF, Miller D, Patton JT, Inohara N, Nunez G: Critical role for cryopyrin/Nalp3 in activation of caspase-1 in response to viral infection and double-stranded RNA. J Biol Chem 2006;281:36560–36568.
- Franchi L, Kanneganti TD, Dubyak GR, Nunez G: Differential requirement of P2X7 receptor and intracellular K+ for caspase-1 activation induced by intracellular and extracellular bacteria. J Biol Chem 2007;282:18810–18818.
- Petrilli V, Papin S, Dostert C, Mayor A, Martinon F, Tschopp J: Activation of the NALP3 inflammasome is triggered by low intracellular potassium concentration. Cell Death Differ 2007;14:1583–1589.
- Hornung V, Bauernfeind F, Halle A, Samstad EO, Kono H, Rock KL, Fitzgerald KA, Latz E: Silica crystals and aluminum salts activate the NALP3 inflammasome through phagosomal destabilization. Nat Immunol 2008;9:847–856.
- Jones JD, Dangl JL: The plant immune system. Nature 2006;444:323–329.
- Chisholm ST, Coaker G, Day B, Staskawicz BJ: Host-microbe interactions: shaping the evolution of the plant immune response. Cell 2006;124:803–814.
Dr. Gabriel Núñez
Department of Pathology and Comprehensive Cancer Center
University of Michigan Medical School
1500 E. Medical Center Drive, Ann Arbor, MI 48109 (USA)
Tel. +1 734 764 8514, Fax +1 734 647 9654, E-Mail firstname.lastname@example.org
Received: March 24, 2009
Accepted after revision: April 30, 2009
Published online: July 1, 2009
Number of Print Pages : 11
Number of Figures : 7, Number of Tables : 0, Number of References : 40
Journal of Innate Immunity
Vol. 1, No. 6, Year 2009 (Cover Date: October 2009)
Journal Editor: Herwald H. (Lund), Egesten A. (Lund)
ISSN: 1662-811X (Print), eISSN: 1662-8128 (Online)
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