title: Prophenoloxidase activation and antimicrobial peptide expression induced by the recombinant microbe binding protein of Manduca sexta.
abstract: Manduca sexta microbe binding protein (MBP) is a member of the β-1,3-glucanase-related protein superfamily that includes Gram-negative bacteria-binding proteins (GNBPs), β-1,3-glucan recognition proteins (βGRPs), and β-1,3-glucanases. Our previous and current studies showed that the purified MBP from baculovirus-infected insect cells had stimulated prophenoloxidase (proPO) activation in the hemolymph of naïve and immune challenged larvae and that supplementation of the exogenous MBP and peptidoglycans (PGs) had caused synergistic increases in PO activity. To explore the underlying mechanism, we separated by SDS-PAGE naïve and induced larval plasma treated with buffer or MBP and detected on immunoblots changes in intensity and/or mobility of hemolymph (serine) proteases [HP14, HP21, HP6, HP8, proPO-activating proteases (PAPs) 1-3] and their homologs (SPH1, SPH2). In a nickel pull-down assay, we observed association of MBP with proHP14 (slightly), βGRP2, PG recognition protein-1 (PGRP1, indirectly), SPH1, SPH2, and proPO2. Further experiments indicated that diaminopimelic acid (DAP) or Lys PG, MBP, PGRP1, and proHP14 together trigger the proPO activation system in a Ca2+-dependent manner. Injection of the recombinant MBP into the 5th instar naïve larvae significantly induced the expression of several antimicrobial peptide genes, revealing a possible link between HP14 and immune signal transduction. Together, these results suggest that the recognition of Gram-negative or -positive bacteria via their PGs induces the melanization and Toll pathways in M. sexta.
authors.affiliation: Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater, OK 74078, United States.
authors.forename: Yang
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater, OK 74078, United States. Electronic address: haobo.jiang@okstate.edu.
authors.forename: Haobo
authors.initials: H
authors.lastname: Jiang
mesh_terms: D000249:Adenosine Monophosphate / Q000378:metabolism*; D000818:Animals; D004156:Catechol Oxidase / Q000378:metabolism; D004792:Enzyme Precursors / Q000378:metabolism; D006458:Hemolymph / Q000378:metabolism; D019476:Insect Proteins / Q000502:physiology*; D018402:Manduca / Q000276:immunology* / Q000378:metabolism
chemical_list: D004792:Enzyme Precursors; D019476:Insect Proteins; D000249:Adenosine Monophosphate; C038451:pro-phenoloxidase; D004156:Catechol Oxidase
keywords: Hemolymph protease system; Insect immunity; Lys- or DAP-type peptidoglycan; Melanization; Pattern recognition; Toll pathway