TOR signaling is required for amino acid stimulation of early trypsin protein synthesis in the midgut of Aedes aegypti mosquitoes

Blood meal digestion in mosquitoes occurs in two phases, an early phase that is translationally regulated, and a late phase that is transcriptionally regulated. Early trypsin is a well-characterized serine endoprotease that is representative of other early phase proteases in the midgut that are only synthesized after feeding. Since the kinase Target of Rapamycin (TOR) has been implicated as a nutrient sensor in other systems, including the mosquito fat body, we tested if TOR signaling is involved in early trypsin protein synthesis in the mosquito midgut in response to feeding. We found that ingestion of an amino acid meal by female mosquitoes induces early trypsin protein synthesis, coincident with phosphorylation of two known TOR target proteins, p70S6 kinase (S6K) and the translational repressor 4E-Binding Protein (4E-BP). Moreover, in vitro culturing of midguts from unfed mosquitoes led to amino acid-dependent phosphorylation of S6K and 4E-BP which could be blocked by treatment with rapamycin, a TOR-specific inhibitor. Lastly, by injecting mosquitoes with TOR double stranded RNA (dsRNA) or rapamycin, we demonstrated that TOR signaling was required in vivo for both phosphorylation of S6K and 4E-BP in the midgut, and for translation of early trypsin mRNA in response to amino acid feeding. It may be possible to target the TOR signaling pathway in the midgut to inhibit blood meal digestion, and thereby, decrease fecundity and the spread of mosquito borne diseases.