Hemocyte surface phenoloxidase (PO) and immune response to lipopolysaccharide (LPS) in Ceratitis capitata.
Bacterial lipopolysaccharide (LPS) attachment at the hemocyte surface is based on the crosslinking of surface associated p47 to LPS, via the intermediacy of tyrosine derivatives generated by the action of phenoloxidase (PO). This attachment is an initial step for LPS internalization from hemocytes (Charalambidis et al., 1996). The results presented clearly show the critical role of hemocyte associated PO activity in the above processes. Biochemical and immunofluorescent analysis demonstrated unambiguously the presence of prophenoloxidase (proPO) on the hemocyte surface. The cell-surface expression of proPO appeared to be LPS-independent, whereas its activation was LPS-dependent. The activation of cell surface proPO involves a limited proteolysis, since upon activation with chymotrypsin proPO is converted to a set of smaller molecular weight proteins with PO activity. The activation appears to be due to enzyme activators, serine proteases, released upon LPS-stimulation. This hypothesis was supported from the activation of membrane proPO by the culture medium of hemocytes which have been triggered with LPS. In addition, proPO, activation was abolished by inhibitors of secretion and PMSF. The release of proPO activators upon LPS-stimulation is mediated via protein tyrosine phosphorylation, as genistein inhibited proPO activation, a situation similar to that reported by us for the release of the effector protein p47 (Charalambidis et al., 1995). The LPS-stimulated activation of cell-surface proPO is a prerequisite for LPS (either cell associated or cell free) internalization, as judged by the resistance of LPS binding to dissociation by proteinase K.
Charalambidis
N D
ND
Department of Biology, University of Patras, Greece.
Foukas
L C
LC
Zervas
C G
CG
Marmaras
V J
VJ
D000818:Animals; D004175:Diptera / Q000201:enzymology* / Q000276:immunology; D005089:Exocytosis / Q000276:immunology; D006434:Hemocytes / Q000201:enzymology* / Q000276:immunology; D008070:Lipopolysaccharides / Q000276:immunology*; D014442:Monophenol Monooxygenase / Q000276:immunology*; D010766:Phosphorylation; D011498:Protein Precursors / Q000276:immunology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors
D008070:Lipopolysaccharides; D011498:Protein Precursors; D014442:Monophenol Monooxygenase; D011505:Protein-Tyrosine Kinases