AgMESH, a peritrophic matrix-associated protein embedded in Anopheles gambiae melanotic capsules modulates malaria parasite infection

Melanins are structurally complex pigments produced by organisms in all domains of life from bacteria to animals, including humans. In insects, melanins are essential for survival and have key roles in cuticle sclerotization, wound healing and innate immunity. In this study, we used a diverse set of molecular, biochemical, and imaging approaches to characterize mosquito melanin involved in innate immune defense (melanotic capsules ). We observed that melanotic capsules enclosing Plasmodium berghei ookinetes were composed of an acid-resistant and highly hydrophobic material with granular appearance, which are characteristic properties of melanins. Spectroscopical analyses reveal chemical signatures of eumelanins and pheomelanin. Furthermore, using an enrichment approach for the proteomic analysis, we identified a set of 14 of acid-resistant mosquito proteins embedded within the melanin matrix possibly related to an anti- Plasmodium response. Among these, Ag Mesh, a highly conserved protein among insect species that contains domains suggesting a role in immune recognition and function. We targeted Ag Mesh for further study using a RNAi-based gene silencing approach in mosquitos and challenged them with two Plasmodium spp. Surprisingly, Ag Mesh gene silencing in mosquitos was associated with reduced parasite infection, implying an important role in facilitating vector infection by Plasmodium spp. Our results provide a new approach to study aspects of insect melanogenesis that revealed proteins associated with melanotic capsule, one of which was strongly implicated in the pathogenesis of Plasmodium spp. mosquito infection. Given the conservation of Ag Mesh among disease-transmitting insect vector species, future analysis of this protein could provide fertile ground for the identification of strategies that block transmission of disease to humans.