The Role Of BmPGRP-S5 In Melanization And Cell Phagocytosis

Peptidoglycan recognition proteins（PGRPs）,as a a very important class of pattern recognition receptors（PRRs）,are widely present in insects.PGRPs can specifically recognize peptidoglycan（PGN）of bacterial cell wall,which triggers the immune response to foreign microorganisms.PGRPs are usually found as family and divided into L-type PGRP and S-type PGRP according to length of their 5’-untranslated region and transcriptional type.13 PGRPs has been identified in the model insect Drosophila melanogaster.And they are involved in the humoral immunity and cellular immunity.12 BmPGRPs,including 6S-types and 6 L-types,were found in the silkworm Bombyx mori.Though BmPGRPs were predicted to be involved in silkworm immunity,structures and functions of them are still unclear.In this study,the constructed recombinant vector BmPGRP-S5-pIEx-4 was transfected into Drosophila embryonic S2 cells to obtain S2 cell line stably expressing BmPGRP-S5.Then a large number of S2 cells were cultured,and the secreted BmPGRP-S5 protein was purified by Ni-NTA,and it’s functions were investigated.The results of the following studies were obtained:（1）The S2 cell line stably expressing BmPGRP-S5 was obtained and recombinant BmPGRP-S5 was purified successfully.（2）BmPGRP-S5 showed weaker antibacterial activities to E.coli、S.aureus,and B.megaterium,but the antibacterial effect to B.megaterium was significantly enhanced in the presence of Zn²⁺.（3）BmPGRP-S5 protein was combined with the cell wall components of E.coli,S.aureus and Bacillus subtilis.The ELISA results showed that BmPGRP-S5 had stronger binding ability to Lys-PGN of S.aureus,LTA of Bacillus subtilis,and Dap-PGN of E.coli,than to TLA of S.aureus and LPS of E.coli,and B.subtilis.（4）The effect of BmPGRPs on the melanization of the silkworm revealed that the BmPGRP-S5 alone could activate the melanization reaction.After adding the cell wall components of E.coli,S.aureus and B.megaterium,the activation of melanization by BmPGRP-S5 was faster.（5）Results of the phagocytosis of hemocytes to E.coli,S.aureus,and B.megaterium revealed that BmPGRP-S5 could promote the phagocytosis of hemocytes to E.coli and S.aureus.This study will help to clarify the identification mechanism of cell phagocytosis by foreign microorganisms.It also provides a new target reference for the further search for the development of new bactericidal drugs.