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 pheno loxidase (PO), This attachment is an initial step for LPS internalizat ion 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 demons trated unambiguously the presence of prophenoloxidase (proPO) on the h emocyte surface, The cell-surface expression of proPO appeared to be L PS-independent, whereas its activation was LPS-dependent. The activati on of cell surface proPO involves a limited proteolysis, since upon ac tivation,vith chymotrypsin proPO is converted to a set of smaller mole cular weight proteins with PO activity, The activation appears to be d ue to enzyme activators, serine proteases, released upon LPS-stimulati on, This hypothesis was supported from the activation of membrane proP O by the culture medium of hemocytes which have been triggered with LP S, In addition, proPO, activation was abolished by inhibitors of secre tion 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 b y the resistance of LPS binding to dissociation by proteinase K. Copyr ight (C) 1996 Elsevier Science Ltd