Proteolytic Cascade for the Activation of the Insect Toll Pathway Induced by the Fungal Cell Wall Component

The insect Toll signaling pathway is activated upon recognition of Gram-positive bacteria and fungi, resulting in the expression of antimicrobial peptides via NF-κB-like transcription factor. This activation is mediated by a serine protease cascade leading to the processing of Spätzle, which generates the functional ligand of the Toll receptor. Recently, we identified three serine proteases mediating Toll pathway activation induced by lysine-type peptidoglycan of Gram-positive bacteria. However, the identities of the downstream serine protease components of Gram-negative-binding protein 3 (GNBP3), a receptor for a major cell wall component β-1,3-glucan of fungi, and their order of activation have not been characterized yet. Here, we identified three serine proteases that are required for Toll activation by β-1,3-glucan in the larvae of a large beetle, Tenebrio molitor. The first one is a modular serine protease functioning immediately downstream of GNBP3 that proteolytically activates the second one, a Spätzle-processing enzyme-activating enzyme that in turn activates the third serine protease, a Spätzle-processing enzyme. The active form of Spätzle-processing enzyme then cleaves Spätzle into the processed Spätzle as Toll ligand. In addition, we show that injection of β-1,3-glucan into Tenebrio larvae induces production of two antimicrobial peptides, Tenecin 1 and Tenecin 2, which are also inducible by injection of the active form of Spätzle-processing enzyme-activating enzyme or processed Spätzle. These results demonstrate a three-step proteolytic cascade essential for the Toll pathway activation by fungal β-1,3-glucan in Tenebrio larvae, which is shared with lysine-type peptidoglycan-induced Toll pathway activation. The insect Toll signaling pathway is activated upon recognition of Gram-positive bacteria and fungi, resulting in the expression of antimicrobial peptides via NF-κB-like transcription factor. This activation is mediated by a serine protease cascade leading to the processing of Spätzle, which generates the functional ligand of the Toll receptor. Recently, we identified three serine proteases mediating Toll pathway activation induced by lysine-type peptidoglycan of Gram-positive bacteria. However, the identities of the downstream serine protease components of Gram-negative-binding protein 3 (GNBP3), a receptor for a major cell wall component β-1,3-glucan of fungi, and their order of activation have not been characterized yet. Here, we identified three serine proteases that are required for Toll activation by β-1,3-glucan in the larvae of a large beetle, Tenebrio molitor. The first one is a modular serine protease functioning immediately downstream of GNBP3 that proteolytically activates the second one, a Spätzle-processing enzyme-activating enzyme that in turn activates the third serine protease, a Spätzle-processing enzyme. The active form of Spätzle-processing enzyme then cleaves Spätzle into the processed Spätzle as Toll ligand. In addition, we show that injection of β-1,3-glucan into Tenebrio larvae induces production of two antimicrobial peptides, Tenecin 1 and Tenecin 2, which are also inducible by injection of the active form of Spätzle-processing enzyme-activating enzyme or processed Spätzle. These results demonstrate a three-step proteolytic cascade essential for the Toll pathway activation by fungal β-1,3-glucan in Tenebrio larvae, which is shared with lysine-type peptidoglycan-induced Toll pathway activation. Innate immunity is a crucial host defense mechanism against microbial infection in all animals (1.Hoffmann J.A. Kafatos F.C. Janeway C.A. Ezekowitz R.A. Science. 1999; 284: 1313-1318Crossref PubMed Scopus (2137) Google Scholar, 2.Medzhitov R. Janeway Jr., C.A. Cell. 1997; 91: 295-298Abstract Full Text Full Text PDF PubMed Scopus (1952) Google Scholar, 3.Akira S. Curr. Opin. Immunol. 2003; 15: 5-11Crossref PubMed Scopus (471) Google Scholar). A group of germ line-encoded receptors and soluble proteins recognizes infectious microbes by sensing specific molecules called pathogen-associated molecular patterns (PAMPs), such as lipopolysaccharides in Gram-negative bacteria, peptidoglycans (PGs) 2The abbreviations used are: PGpeptidoglycanAMPantimicrobial peptidePGRPPG recognition proteinGNBPGram-negative-binding proteinSPserine proteaseMSPmodular serine proteaseSPESpätzle-processing enzymeSAESPE-activating enzymei.d.inner diameter.2The abbreviations used are: PGpeptidoglycanAMPantimicrobial peptidePGRPPG recognition proteinGNBPGram-negative-binding proteinSPserine proteaseMSPmodular serine proteaseSPESpätzle-processing enzymeSAESPE-activating enzymei.d.inner diameter. in Gram-positive bacteria, and β-1,3-glucans in fungi (4.Janeway Jr., C.A. Cold Spring Harb. Symp. Quant. Biol. 1989; 54: 1-13Crossref PubMed Google Scholar, 5.Lemaitre B. Nicolas E. Michaut L. Reichhart J.M. Hoffmann J.A. Cell. 1996; 86: 973-983Abstract Full Text Full Text PDF PubMed Scopus (2955) Google Scholar, 6.Medzhitov R. Janeway Jr., C.A. Science. 2002; 296: 298-300Crossref PubMed Scopus (1639) Google Scholar, 7.Ferrandon D. Imler J.L. Hetru C. Hoffmann J.A. Nat. Rev. Immunol. 2007; 7: 862-874Crossref PubMed Scopus (621) Google Scholar). The Drosophila Toll signaling pathway is activated upon recognition of lysine (Lys)-type PG, which is found in Gram-positive bacteria and β-1,3-glucan, a major component of the fungus cell wall (8.Levashina E.A. Langley E. Green C. Gubb D. Ashburner M. Hoffmann J.A. Reichhart J.M. Science. 1999; 285: 1917-1919Crossref PubMed Scopus (368) Google Scholar, 9.El Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). In contrast, the immune deficiency (Imd) pathway is activated primarily after recognition of diaminopimelic acid (DAP)-type PG, a form of PG found in Gram-negative bacteria and Bacillus spp (10.Leulier F. Parquet C. Pili-Floury S. Ryu J.H. Caroff M. Lee W.J. Mengin-Lecreulx D. Lemaitre B. Nat. Immunol. 2003; 4: 478-484Crossref PubMed Scopus (447) Google Scholar). Both pathways lead to the expression of antimicrobial peptides (AMPs) via NF-κB-like transcription factors (11.Royet J. Reichhart J.M. Hoffmann J.A. Curr. Opin. Immunol. 2005; 17: 11-17Crossref PubMed Scopus (135) Google Scholar, 12.Cherry S. Silverman N. Nat. Immunol. 2006; 7: 911-917Crossref PubMed Scopus (178) Google Scholar, 13.Lemaitre B. Hoffmann J. Annu. Rev. Immunol. 2007; 25: 697-743Crossref PubMed Scopus (2301) Google Scholar).Elegant genetic studies in Drosophila demonstrated that Lys-type PG is recognized by a complex comprised of the PG recognition protein (PGRP)-SA (14.Michel T. Reichhart J.M. Hoffmann J.A. Royet J. Nature. 2001; 414: 756-759Crossref PubMed Scopus (604) Google Scholar), PGRP-SD (15.Bischoff V. Vignal C. Boneca I.G. Michel T. Hoffmann J.A. Royet J. Nat. Immunol. 2004; 5: 1175-1180Crossref PubMed Scopus (199) Google Scholar), and the Gram-negative-binding protein 1 (GNBP1) (16.Gobert V. Gottar M. Matskevich A.A. Rutschmann S. Royet J. Belvin M. Hoffmann J.A. Ferrandon D. Science. 2003; 302: 2126-2130Crossref PubMed Scopus (297) Google Scholar, 17.Pili-Floury S. Leulier F. Takahashi K. Saigo K. Samain E. Ueda R. Lemaitre B. J. Biol. Chem. 2004; 279: 12848-12853Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar), while β-1,3-glucan from yeast is recognized by GNBP3 (18.Gottar M. Gobert V. Matskevich A.A. Reichhart J.M. Wang C. Butt T.M. Belvin M. Hoffmann J.A. Ferrandon D. Cell. 2006; 127: 1425-1437Abstract Full Text Full Text PDF PubMed Scopus (311) Google Scholar). Both the PGRP-SA/GNBP1 complex and GNBP3 are believed to mediate the activation of a serine protease cascade that ultimately leads to the cleavage of pro-Spätzle into processed Spätzle (5.Lemaitre B. Nicolas E. Michaut L. Reichhart J.M. Hoffmann J.A. Cell. 1996; 86: 973-983Abstract Full Text Full Text PDF PubMed Scopus (2955) Google Scholar, 19.Weber A.N. Tauszig-Delamasure S. Hoffmann J.A. Lelièvre E. Gascan H. Ray K.P. Morse M.A. Imler J.L. Gay N.J. Nat. Immunol. 2003; 4: 794-800Crossref PubMed Scopus (329) Google Scholar, 20.Gay N.J. Gangloff M. Annu. Rev. Biochem. 2007; 76: 141-165Crossref PubMed Scopus (505) Google Scholar). Drosophila Spätzle-processing enzyme (SPE), serine protease with a been identified as the serine protease that cleaves pro-Spätzle N. Lemaitre B. M. S. C. M. Lee W.J. Cell. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar, S. Takahashi K. Lee W.J. Ueda R. Lemaitre B. Curr. Biol. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). Spätzle as a ligand for the cell receptor and induces the production of by the (8.Levashina E.A. Langley E. Green C. Gubb D. Ashburner M. Hoffmann J.A. Reichhart J.M. Science. 1999; 285: 1917-1919Crossref PubMed Scopus (368) Google Scholar, 19.Weber A.N. Tauszig-Delamasure S. Hoffmann J.A. Lelièvre E. Gascan H. Ray K.P. Morse M.A. Imler J.L. Gay N.J. Nat. Immunol. 2003; 4: 794-800Crossref PubMed Scopus (329) Google Scholar, 20.Gay N.J. Gangloff M. Annu. Rev. Biochem. 2007; 76: 141-165Crossref PubMed Scopus (505) Google Scholar). protease in and from recognition receptors in the insect and lead to activation of the signaling pathway C. Biol. 1999; 9: Full Text Full Text PDF PubMed Scopus Google Scholar, Lee W.J. 2008; PubMed Google Scholar). the activation of serine proteases and to the B. Hoffmann J. Annu. Rev. Immunol. 2007; 25: 697-743Crossref PubMed Scopus (2301) Google Scholar, T. Nat. Rev. Immunol. 2002; PubMed Scopus Google Scholar). The of recognition host defense in which immune (1.Hoffmann J.A. Kafatos F.C. Janeway C.A. Ezekowitz R.A. Science. 1999; 284: 1313-1318Crossref PubMed Scopus (2137) Google we the serine protease cascade that the Toll pathway larvae of the beetle, Tenebrio H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar, H. Lee H. R. J. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). This large insect to large of to serine demonstrated that the recognition of Lys-type PG by the PGRP-SA/GNBP1 complex activates Spätzle the activation of three Tenebrio serine modular serine protease Spätzle-processing enzyme-activating enzyme and H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google activation mechanism of the Toll pathway by β-1,3-glucan is characterized in A Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google that Drosophila as a serine protease, which Gram-positive bacteria and fungi via the recognition receptors Drosophila PGRP-SA/GNBP1 and that the Drosophila protease, to specific for fungal in the Toll pathway (18.Gottar M. Gobert V. Matskevich A.A. Reichhart J.M. Wang C. Butt T.M. Belvin M. Hoffmann J.A. Ferrandon D. Cell. 2006; 127: 1425-1437Abstract Full Text Full Text PDF PubMed Scopus (311) Google Scholar, N. Hoffmann J.A. Reichhart J.M. Science. 2002; PubMed Scopus Google Scholar), also required for the sensing of proteases by bacteria in the Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). Drosophila and proteases are to serine proteases that fungal recognition to pro-Spätzle Drosophila Toll pathway the of serine proteases have not been the and of the downstream of GNBP3 or the serine of Spätzle that in the Toll signaling pathway are essential for the host defense against fungal and the sensing of fungal we have identified the serine protease that immediately downstream of Tenebrio in we as to the Tenebrio β-1,3-glucan recognition is to Spätzle. identified three Tenebrio serine and that in a cascade β-1,3-glucan recognition by GNBP3 to pro-Spätzle activation. we identified two that induced upon injection of β-1,3-glucan, activated or a processed Spätzle into Tenebrio larvae, that of the Toll we three the β-1,3-glucan recognition signaling pathway in T. show Tenebrio downstream of the β-1,3-glucan recognition is via the activation of the three Tenebrio serine and and that results in the processing of pro-Spätzle to and the activation of the cascade the expression of two Tenecin 1 and Tenecin results demonstrate that the Tenebrio cascade is essential in the recognition of fungi and in the activation of antimicrobial defense with H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar), a in which Lys-type PG and β-1,3-glucan a of three serine protease This three-step proteolytic processing of the pro-Spätzle active Spätzle, which then to the Toll resulting in the of expression in the Tenebrio of the three serine proteases to activation of the Tenebrio Toll The enzyme is serine protease with cleavage and a serine protease is a serine protease with cleavage and a serine protease and is a protein with cleavage and a serine protease These three serine protease the of the proteolytic serine protease cascade and cleavage of serine protease genetic studies in or have demonstrated that Drosophila serine such as and of the Drosophila activation of Toll S. Takahashi K. Lee W.J. Ueda R. Lemaitre B. Curr. Biol. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). Chamy Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google demonstrated that Drosophila as a signaling component required for the of fungi by GNBP3 and the sensing of Gram-positive bacteria by that Drosophila serine protease, to specific for fungal required for the sensing of proteases that are by fungi and bacteria Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). This that Drosophila a proteolytic cascade activated by factors in the However, Drosophila and not show with Tenebrio which as serine protease to Tenebrio GNBP3 and PGRP-SA/GNBP1 These results the of in the proteolytic of Toll in Tenebrio and In to of serine the expression of Tenebrio are also from of Drosophila The Tenebrio Tenecin 1 induced in of the Tenebrio larvae, while Drosophila are by the the in the of the serine protease in the of and in the of Drosophila and Tenebrio to specific or to in their and in the used by to microbial infection the of the immune in we that the activated Tenebrio the Tenebrio and serine protease 1 to their to active complex H. Lee H. R. J. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). This of a and a serine protease the of bacteria, and a H. Lee H. R. J. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). Tenebrio also in the activation of the Toll pathway by GNBP3 upon β-1,3-glucan sensing and by upon Lys-type PG results that the and the Toll pathway immune are a serine protease cascade for the of two major immune studies the molecular mechanism the Toll pathway by a in which Lys-type PG and β-1,3-glucan recognition a proteolytic cascade three serine protease that are This three-step proteolytic cascade leads to the of Spätzle and the activation of the Toll signaling that the of A of two also the of to bacteria and fungi in and and Innate immunity is a crucial host defense mechanism against microbial infection in all animals (1.Hoffmann J.A. Kafatos F.C. Janeway C.A. Ezekowitz R.A. Science. 1999; 284: 1313-1318Crossref PubMed Scopus (2137) Google Scholar, 2.Medzhitov R. Janeway Jr., C.A. Cell. 1997; 91: 295-298Abstract Full Text Full Text PDF PubMed Scopus (1952) Google Scholar, 3.Akira S. Curr. Opin. Immunol. 2003; 15: 5-11Crossref PubMed Scopus (471) Google Scholar). A group of germ line-encoded receptors and soluble proteins recognizes infectious microbes by sensing specific molecules called pathogen-associated molecular patterns (PAMPs), such as lipopolysaccharides in Gram-negative bacteria, peptidoglycans (PGs) 2The abbreviations used are: PGpeptidoglycanAMPantimicrobial peptidePGRPPG recognition proteinGNBPGram-negative-binding proteinSPserine proteaseMSPmodular serine proteaseSPESpätzle-processing enzymeSAESPE-activating enzymei.d.inner diameter.2The abbreviations used are: PGpeptidoglycanAMPantimicrobial peptidePGRPPG recognition proteinGNBPGram-negative-binding proteinSPserine proteaseMSPmodular serine proteaseSPESpätzle-processing enzymeSAESPE-activating enzymei.d.inner diameter. in Gram-positive bacteria, and β-1,3-glucans in fungi (4.Janeway Jr., C.A. Cold Spring Harb. Symp. Quant. Biol. 1989; 54: 1-13Crossref PubMed Google Scholar, 5.Lemaitre B. Nicolas E. Michaut L. Reichhart J.M. Hoffmann J.A. Cell. 1996; 86: 973-983Abstract Full Text Full Text PDF PubMed Scopus (2955) Google Scholar, 6.Medzhitov R. Janeway Jr., C.A. Science. 2002; 296: 298-300Crossref PubMed Scopus (1639) Google Scholar, 7.Ferrandon D. Imler J.L. Hetru C. Hoffmann J.A. Nat. Rev. Immunol. 2007; 7: 862-874Crossref PubMed Scopus (621) Google Scholar). The Drosophila Toll signaling pathway is activated upon recognition of lysine (Lys)-type PG, which is found in Gram-positive bacteria and β-1,3-glucan, a major component of the fungus cell wall (8.Levashina E.A. Langley E. Green C. Gubb D. Ashburner M. Hoffmann J.A. Reichhart J.M. Science. 1999; 285: 1917-1919Crossref PubMed Scopus (368) Google Scholar, 9.El Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). In contrast, the immune deficiency (Imd) pathway is activated primarily after recognition of diaminopimelic acid (DAP)-type PG, a form of PG found in Gram-negative bacteria and Bacillus spp (10.Leulier F. Parquet C. Pili-Floury S. Ryu J.H. Caroff M. Lee W.J. Mengin-Lecreulx D. Lemaitre B. Nat. Immunol. 2003; 4: 478-484Crossref PubMed Scopus (447) Google Scholar). Both pathways lead to the expression of antimicrobial peptides (AMPs) via NF-κB-like transcription factors (11.Royet J. Reichhart J.M. Hoffmann J.A. Curr. Opin. Immunol. 2005; 17: 11-17Crossref PubMed Scopus (135) Google Scholar, 12.Cherry S. Silverman N. Nat. Immunol. 2006; 7: 911-917Crossref PubMed Scopus (178) Google Scholar, 13.Lemaitre B. Hoffmann J. Annu. Rev. Immunol. 2007; 25: 697-743Crossref PubMed Scopus (2301) Google Scholar). peptidoglycan antimicrobial PG recognition protein Gram-negative-binding protein serine protease modular serine protease Spätzle-processing enzyme enzyme diameter. peptidoglycan antimicrobial PG recognition protein Gram-negative-binding protein serine protease modular serine protease Spätzle-processing enzyme enzyme diameter. genetic studies in Drosophila demonstrated that Lys-type PG is recognized by a complex comprised of the PG recognition protein (PGRP)-SA (14.Michel T. Reichhart J.M. Hoffmann J.A. Royet J. Nature. 2001; 414: 756-759Crossref PubMed Scopus (604) Google Scholar), PGRP-SD (15.Bischoff V. Vignal C. Boneca I.G. Michel T. Hoffmann J.A. Royet J. Nat. Immunol. 2004; 5: 1175-1180Crossref PubMed Scopus (199) Google Scholar), and the Gram-negative-binding protein 1 (GNBP1) (16.Gobert V. Gottar M. Matskevich A.A. Rutschmann S. Royet J. Belvin M. Hoffmann J.A. Ferrandon D. Science. 2003; 302: 2126-2130Crossref PubMed Scopus (297) Google Scholar, 17.Pili-Floury S. Leulier F. Takahashi K. Saigo K. Samain E. Ueda R. Lemaitre B. J. Biol. Chem. 2004; 279: 12848-12853Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar), while β-1,3-glucan from yeast is recognized by GNBP3 (18.Gottar M. Gobert V. Matskevich A.A. Reichhart J.M. Wang C. Butt T.M. Belvin M. Hoffmann J.A. Ferrandon D. Cell. 2006; 127: 1425-1437Abstract Full Text Full Text PDF PubMed Scopus (311) Google Scholar). Both the PGRP-SA/GNBP1 complex and GNBP3 are believed to mediate the activation of a serine protease cascade that ultimately leads to the cleavage of pro-Spätzle into processed Spätzle (5.Lemaitre B. Nicolas E. Michaut L. Reichhart J.M. Hoffmann J.A. Cell. 1996; 86: 973-983Abstract Full Text Full Text PDF PubMed Scopus (2955) Google Scholar, 19.Weber A.N. Tauszig-Delamasure S. Hoffmann J.A. Lelièvre E. Gascan H. Ray K.P. Morse M.A. Imler J.L. Gay N.J. Nat. Immunol. 2003; 4: 794-800Crossref PubMed Scopus (329) Google Scholar, 20.Gay N.J. Gangloff M. Annu. Rev. Biochem. 2007; 76: 141-165Crossref PubMed Scopus (505) Google Scholar). Drosophila Spätzle-processing enzyme (SPE), serine protease with a been identified as the serine protease that cleaves pro-Spätzle N. Lemaitre B. M. S. C. M. Lee W.J. Cell. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar, S. Takahashi K. Lee W.J. Ueda R. Lemaitre B. Curr. Biol. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). Spätzle as a ligand for the cell receptor and induces the production of by the (8.Levashina E.A. Langley E. Green C. Gubb D. Ashburner M. Hoffmann J.A. Reichhart J.M. Science. 1999; 285: 1917-1919Crossref PubMed Scopus (368) Google Scholar, 19.Weber A.N. Tauszig-Delamasure S. Hoffmann J.A. Lelièvre E. Gascan H. Ray K.P. Morse M.A. Imler J.L. Gay N.J. Nat. Immunol. 2003; 4: 794-800Crossref PubMed Scopus (329) Google Scholar, 20.Gay N.J. Gangloff M. Annu. Rev. Biochem. 2007; 76: 141-165Crossref PubMed Scopus (505) Google Scholar). protease in and from recognition receptors in the insect and lead to activation of the signaling pathway C. Biol. 1999; 9: Full Text Full Text PDF PubMed Scopus Google Scholar, Lee W.J. 2008; PubMed Google Scholar). the activation of serine proteases and to the B. Hoffmann J. Annu. Rev. Immunol. 2007; 25: 697-743Crossref PubMed Scopus (2301) Google Scholar, T. Nat. Rev. Immunol. 2002; PubMed Scopus Google Scholar). The of recognition host defense in which immune (1.Hoffmann J.A. Kafatos F.C. Janeway C.A. Ezekowitz R.A. Science. 1999; 284: 1313-1318Crossref PubMed Scopus (2137) Google Scholar). Recently, we the serine protease cascade that the Toll pathway larvae of the beetle, Tenebrio H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar, H. Lee H. R. J. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). This large insect to large of to serine demonstrated that the recognition of Lys-type PG by the PGRP-SA/GNBP1 complex activates Spätzle the activation of three Tenebrio serine modular serine protease Spätzle-processing enzyme-activating enzyme and H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). The activation mechanism of the Toll pathway by β-1,3-glucan is characterized in A Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google that Drosophila as a serine protease, which Gram-positive bacteria and fungi via the recognition receptors Drosophila PGRP-SA/GNBP1 and that the Drosophila protease, to specific for fungal in the Toll pathway (18.Gottar M. Gobert V. Matskevich A.A. Reichhart J.M. Wang C. Butt T.M. Belvin M. Hoffmann J.A. Ferrandon D. Cell. 2006; 127: 1425-1437Abstract Full Text Full Text PDF PubMed Scopus (311) Google Scholar, N. Hoffmann J.A. Reichhart J.M. Science. 2002; PubMed Scopus Google Scholar), also required for the sensing of proteases by bacteria in the Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). Drosophila and proteases are to serine proteases that fungal recognition to pro-Spätzle Drosophila Toll pathway the of serine proteases have not been the and of the downstream of GNBP3 or the serine of Spätzle that in the Toll signaling pathway are essential for the host defense against fungal and the sensing of fungal Here, we have identified the serine protease that immediately downstream of Tenebrio in we as to the Tenebrio β-1,3-glucan recognition is to Spätzle. identified three Tenebrio serine and that in a cascade β-1,3-glucan recognition by GNBP3 to pro-Spätzle activation. we identified two that induced upon injection of β-1,3-glucan, activated or a processed Spätzle into Tenebrio larvae, that of the Toll we three the β-1,3-glucan recognition signaling pathway in T. show Tenebrio downstream of the β-1,3-glucan recognition is via the activation of the three Tenebrio serine and and that results in the processing of pro-Spätzle to and the activation of the cascade the expression of two Tenecin 1 and Tenecin results demonstrate that the Tenebrio cascade is essential in the recognition of fungi and in the activation of antimicrobial defense with H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar), a in which Lys-type PG and β-1,3-glucan a of three serine protease This three-step proteolytic processing of the pro-Spätzle active Spätzle, which then to the Toll resulting in the of expression in the Tenebrio of the three serine proteases to activation of the Tenebrio Toll The enzyme is serine protease with cleavage and a serine protease is a serine protease with cleavage and a serine protease and is a protein with cleavage and a serine protease These three serine protease the of the proteolytic serine protease cascade and cleavage of serine protease genetic studies in or have demonstrated that Drosophila serine such as and of the Drosophila activation of Toll S. Takahashi K. Lee W.J. Ueda R. Lemaitre B. Curr. Biol. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). Chamy Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google demonstrated that Drosophila as a signaling component required for the of fungi by GNBP3 and the sensing of Gram-positive bacteria by that Drosophila serine protease, to specific for fungal required for the sensing of proteases that are by fungi and bacteria Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). This that Drosophila a proteolytic cascade activated by factors in the However, Drosophila and not show with Tenebrio which as serine protease to Tenebrio GNBP3 and PGRP-SA/GNBP1 These results the of in the proteolytic of Toll in Tenebrio and In to of serine the expression of Tenebrio are also from of Drosophila The Tenebrio Tenecin 1 induced in of the Tenebrio larvae, while Drosophila are by the the in the of the serine protease in the of and in the of Drosophila and Tenebrio to specific or to in their and in the used by to microbial infection the of the immune in we that the activated Tenebrio the Tenebrio and serine protease 1 to their to active complex H. Lee H. R. J. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). This of a and a serine protease the of bacteria, and a H. Lee H. R. J. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). Tenebrio also in the activation of the Toll pathway by GNBP3 upon β-1,3-glucan sensing and by upon Lys-type PG results that the and the Toll pathway immune are a serine protease cascade for the of two major immune studies the molecular mechanism the Toll pathway by a in which Lys-type PG and β-1,3-glucan recognition a proteolytic cascade three serine protease that are This three-step proteolytic cascade leads to the of Spätzle and the activation of the Toll signaling that the of A of two also the of to bacteria and fungi in and and In we three the β-1,3-glucan recognition signaling pathway in T. show Tenebrio downstream of the β-1,3-glucan recognition is via the activation of the three Tenebrio serine and and that results in the processing of pro-Spätzle to and the activation of the cascade the expression of two Tenecin 1 and Tenecin results demonstrate that the Tenebrio cascade is essential in the recognition of fungi and in the activation of antimicrobial defense with H. R. Lee M. K. Lee J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar), a in which Lys-type PG and β-1,3-glucan a of three serine protease This three-step proteolytic processing of the pro-Spätzle active Spätzle, which then to the Toll resulting in the of expression in the Tenebrio of the three serine proteases to activation of the Tenebrio Toll The enzyme is serine protease with cleavage and a serine protease is a serine protease with cleavage and a serine protease and is a protein with cleavage and a serine protease These three serine protease the of the proteolytic serine protease cascade and cleavage of serine protease genetic studies in or have demonstrated that Drosophila serine such as and of the Drosophila activation of Toll S. Takahashi K. Lee W.J. Ueda R. Lemaitre B. Curr. Biol. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). Chamy Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google demonstrated that Drosophila as a signaling component required for the of fungi by GNBP3 and the sensing of Gram-positive bacteria by that Drosophila serine protease, to specific for fungal required for the sensing of proteases that are by fungi and bacteria Chamy L. Leclerc V. Caldelari I. Reichhart J.M. Nat. Immunol. 2008; 9: 1165-1170Crossref PubMed Scopus (145) Google Scholar). 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