Oncopeltus antibacterial peptide 4

In Oncopeltus fasciatus, evidence shown here indicates it is calmodulin (CaM) that activates phospholipase-C (PLC), beginning a signalling pathway necessary for endocytic uptake of yolk precursor molecules. Epithelial cell-produced CaM, transported to oocytes via gap junctions, has been shown to be required for receptor-mediated endocytic uptake of vitellogenins (Vgs, the protein precursors of yolk). To determine if CaM was directly or indirectly stimulating the phospholipase-C (PLC) signalling cascade and thus controlling Vg endocytosis we used a series of molecules known to inactivate various elements of the pathway. W-7 prevents CaM from interacting with other molecules. Neomycin isolates PIP(2) from PLC. U-73122 directly inactivates PLC. 2-APB blocks IP(3) receptors which would otherwise cause release of Ca(2+). Verapamil and CdCl(2) block Ca(2+) release channels. Staurosporin and calphostin are inhibitors of PK-C. 1-Hexadecyl-2-acetyl glycerol (HAG) binds to diacylglycerol (DAG). Through the use of these antagonists we show here that: (1) the activation of phospholipase-C in this system requires CaM. (2) Stimulated phospholipase-C converts PIP(2) into IP(3) and DAG. (3) IP(3) causes increase in cytosolic Ca(2+). (4) DAG and Ca(2+) each stimulate phosphokinase-C, resulting in endocytosis of Vgs.

Pseudomonas aeruginosa and Pseudomonas putida show a profound differential infectivity after inoculation in Oncopeltus fasciatus. Whereas P. putida has no significant impact on nymphs, P. aeruginosa kills all experimental animals within 48 h. Both Pseudomonas species, however, induce the same four hemolymph peptides in O. fasciatus. Also injection of saline solution and injury induced these peptides. In general peptide induction was stronger in nymphs than in adult males. A significantly higher number of nymphs survived a challenge with P. aeruginosa when an immunization with P. putida preceded. The antibacterial properties of the hemolymph were demonstrated in inhibition experiments with P. putida. Two of the four inducible peptides (peptides 1 and 4) could be partially sequenced after Edman degradation and were compared with known antibacterial peptides. Peptide 1, of 15 kDa, showed 47.1% identity with the glycine-rich hemiptericin of Pyrrhocoris apterus. Peptide 4, of 2 kDa, had a 77.8% identity with the proline-rich pyrrhocoricin of P. apterus and a 76.9% identity with metalnikowin 1 of Palomena prasina. Peptides 2 and 3 are also small, with molecular weights of 8 and 5 kDa.