Identification of a cis-regulatory element required for 20-hydroxyecdysone enhancement immune gene expression in Drosophila melanogaster

Diptericin, an antimicrobial peptide, plays a key role in defense against invading microorganisms. Quantitative real time PCR analyses of the mRNA levels of Diptericin gene during Drosophila melanogaster development showed an increase in the expression of Diptericin gene during the late final instar and early pupal stage when ecdysteroid titers increased to regulate the larval-pupal metamorphosis. Deletion analysis of the 3 kb Diptericin promoter region by reporter assays identified a cis-regulatory element, AAGAAAGATCCCCTG, that was necessary for ecdysone enhancement of the peptidoglycan induced Diptericin gene expression. Nuclear proteins extracted from D. melanogaster mbn2 cells treated with peptidoglycan plus 20-hydroxyecdysone (20E) bound to this element. The peptidoglycan induced expressions of the genes coding for four other AMPs (Drs, AttA, Mtk and CecA1) were also enhanced by 20E. MEME/MAST alignment analysis showed that the presence of an eight nucleotide consensus element, [A(AG)(ATG)TCCCC], a region of the identified cis-regulatory element in the promoter regions of these four AMP genes as well. These studies suggest that 20E regulates the expression of AMP genes through a conserved mechanism. Supported by NIH grant GM 070559-04.