Factors functioning in nodule melanization of insects and their mechanisms of accumulation in nodules

Abstract

Nodules consisting of hemocytes and trapped microorganisms are important targets for melanization, which is best known in the insect immune system. We investigated factors functioning in nodule melanization and the mechanism by which these factors congregate in the nodule. BmHP21, BmSPH1 and BmSPH2, Bombyx mori orthologs of Manduca sexta serine protease HP21, serine protease homologs (SPH1 and SPH2), and a prophenoloxidase, BmPO1 were observed as inactive forms in the plasma, but as putatively active forms in the nodule. Production of prophenoloxidase-activating proteinases, BmPAP1 and BmPAP3/PPAE and BmPO1 were confirmed in hemocytes. BmSPH1 and BmSPH2 were observed on trapped bacterial cells in the nodule and were isolated from the surface of bacterial cells incubated with plasma. BmSPH1 and BmSPH2 were found in plasma in complex with a pattern recognition receptor, BmLBP. These data suggest that melanization-regulating factors congregate in nodules through a combination of microorganism-dependent and hemocyte-dependent routes.

Keywords: 4′,6-diamidino-2-phenylindole; B. mori multibinding protein; BmLBP; BmMBP; Bombyx mori; Bombyx mori lipopolysaccharide binding protein; DAPI; Innate immunity; Melanization; Nodule; PAP; PO; PPAE; Prophenoloxidase; ROS; SP; SPH; Serine protease homolog; phenoloxidase; proPO; proPO-activating enzyme; prophenoloxidase; prophenoloxidase-activating proteinase; reactive oxygen species; serine protease; serine protease homolog.