Identification and Functional Analysis of Antifungal Immune Response Genes in Drosophila

Abstract

Essential aspects of the innate immune response to microbial infection appear to be conserved between insects and mammals. Although signaling pathways that activate NF-κB during innate immune responses to various microorganisms have been studied in detail, regulatory mechanisms that control other immune responses to fungal infection require further investigation. To identify new Drosophila genes involved in antifungal immune responses, we selected genes known to be differentially regulated in SL2 cells by microbial cell wall components and tested their roles in antifungal defense using mutant flies. From 130 mutant lines, sixteen mutants exhibited increased sensitivity to fungal infection. Examination of their effects on defense against various types of bacteria and fungi revealed nine genes that are involved specifically in defense against fungal infection. All of these mutants displayed defects in phagocytosis or activation of antimicrobial peptide genes following infection. In some mutants, these immune deficiencies were attributed to defects in hemocyte development and differentiation, while other mutants showed specific defects in immune signaling required for humoral or cellular immune responses. Our results identify a new class of genes involved in antifungal immune responses in Drosophila.

Requirement of Split ends for Epigenetic Regulation of Notch Signal-Dependent Genes during Infection-Induced Hemocyte Differentiation

“…The requirement for spen for proper immune development was also shown by the formation of melanotic tumors in spen GE10359 mutant flies (see Fig. S1 in the supplemental material), as well as by increased proliferation of plasmatocytes, and reduced differentiation of crystal cells (15). Together, these results indicate that Spen is required for Drosophila innate immunity by regulating hemocytes development.…”

“…Spen is required for activation of Notch-dependent genes. The defects that we observed in the spen mutant hemocytes are often seen in association with inappropriate hemocyte development (15). The Notch pathway plays an instructive role in the development of hemocytes, and mammalian homologs of Spen (SHARP and MINT) have been proposed to be negative regulators of the Notch signaling pathway (9,23,34,35).…”

“…In a previous study, we identified a spen allele that showed defects in defense against microbial infection and in wound healing (15). To confirm the requirement of Spen for the innate immune response, we analyzed the survival of flies harboring several spen mutant alleles (spen GE10359 , spen poc361 , and spen 03350 ), as well as of flies of the precise excision derivative (spen rev2 ) of spen GE10359 after fungal infection.…”

“…In a previous study, we showed that spen mutant flies have abnormal numbers of hemocytes, phagocytic defects, and melanotic tumors, pointing to a major defect in spen hemocytes (15). In order to identify the molecular defects in the mutant hemocytes, we collected hemocytes from 200 to 300 third-instar wild-type and mutant larvae and analyzed their transcript profiles using Affymetrix arrays.…”

“…We previously isolated a mutant allele of spen (spen GE10359 ) that results in defective immunity to Beauveria bassiana infection and inappropriate hemocyte development (15). In view of the role of Spen as a key regulator of Notch signaling in Drosophila neural development and mammalian hematopoiesis, the compromised immunity shown in the spen mutant flies appears to reflect defects in the immune cells caused by inappropriate development.…”