Abstract
Malpighian tubules (MT) of Drosophila melanogaster are osmoregulatory organs that maintain the ionic balance and remove toxic substances from the body. Additionally they act as autonomous immune sensing organs, which secrete antimicrobial peptides in response to invading microbial pathogens. We show that the antimicrobial peptides (AMP) diptericin, cecropinA, drosocin and attacinA are constitutively expressed and are regulated in developmental stage specific manner. Their developmental expression begins from 3rd instar larval stage and an immune challenge increases the expression several folds. Spatial variatons in the level of expression along the MT tissue are observed. The mortality of 3rd instar larvae fed on bacterial food is much less than that of the earlier larval stages, coinciding with the onset of innate immunity response in MT. Ectopic expression of AMP imparts better resistance to infection while, loss of function of one of the AMP through directed RNAi reduces host survival after immune challenge. The AMP secreted from the MT exhibit bactericidal activity. Expression of the NF-κB transcription factor, Relish, also coincides with activation of immune responsive genes in MT, demonstrating that immune regulation in MT is under developmental control and is governed by the Imd pathway.
Highlights
Innate immunity is an evolutionarily conserved mechanism in eukaryotes and is the first line of defense which enables the organism to survive different infectious pathogens in a non-specific manner
We have examined expression of four antimicrobial peptides (AMP), diptericin, cecropinA, attacinA and drosocin induced by Gram negative bacteria, and which are activated by the Imd pathway
Taking advantage of a robust survival and physiological activity of Malpighian tubules (MT) in culture medium [35,36] and their capability to respond to LPS challenge [25], we monitored the b-galactosidase activity of Lac-Z reporters under diptericin and cecropinA promoters and green fluorescence in GFP construct under drosocin and attacinA promoters after exposing the isolated MT to LPS challenge
Summary
Innate immunity is an evolutionarily conserved mechanism in eukaryotes and is the first line of defense which enables the organism to survive different infectious pathogens in a non-specific manner. In Drosophila, the innate immunity comprises of activation of humoral response resulting in the production of AMP [4], activation and phagocytosis of pathogens by blood cells, plasmatocytes [5] and melanization by the activation of phenoloxidase pathway [2,3], [6,7,8]. The 20 AMP characterized in Drosophila can be arranged into seven different groups, viz., cecropin, diptericin, attacin, drosocin, defensin, drosomycin and metchnikowin, with distinct but some overlapping specificities [9], [10,11]. Diptericin, cecropin and drosocin are active against Gram negative bacteria, metchnikowin and defensin act against Gram positive bacteria and fungi whereas drosomycin is active only against fungi [12]. Cecropin has been known to have four transcripts (A1, A2, B and C), diptericin has two (A and B) and attacin has four (A, B, C and D)
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