Carbon dioxide (CO(2)) is an important long-range chemosensory cue used by blood-feeding female mosquitoes to find their hosts. The CO(2) receptor in Drosophila melanogaster was previously determined to be a heterodimer comprised of two gustatory receptor (Gr) proteins, DmGr21a and DmGr63a. In the mosquito Aedes aegypti, two putative orthologous genes, AaGr1 and AaGr3, were identified in the genome database, along with an apparent paralogue of AaGr1, AaGr2. In this study, RNA interference (RNAi)-mediated gene knockdown of either AaGr1 or AaGr3 resulted in a loss of CO(2) sensitivity in both male and female mosquitoes, suggesting that these two proteins, like the Drosophila orthologues, function as a heterodimer. RNAi-mediated knockdown of AaGr2 expression had no impact on CO(2) reception. All three Gr genes were expressed in the maxillary palps of both Ae. aegypti and the West Nile virus vector mosquito, Culex pipiens quinquefasciatus. Interestingly, expression of the two CO(2) receptor genes was not equivalent in the two sexes and the implications of differential sex expression of the CO(2) receptor in different species are discussed. The functional identification of the CO(2) receptor in a mosquito could prove invaluable in the strategic design of compounds that disrupt the mosquito's ability to find hosts.
Our data indicate that miR-200b improves PH and decreases the incidence of CDH. Future studies will further exploit this newly discovered prenatal therapy for lung hypoplasia and CDH.
Subcellular trafficking within host cells plays a critical role in viral life cycles, including influenza A virus (IAV). Thus targeting relevant subcellular compartments holds promise for effective intervention to control the impact of influenza infection. Bafilomycin A1 (Baf-A1), when used at relative high concentrations (≥10 nM), inhibits vacuolar ATPase (V-ATPase) and reduces endosome acidification and lysosome number, thus inhibiting IAV replication but promoting host cell cytotoxicity. We tested the hypothesis that much lower doses of Baf-A1 also have anti-IAV activity, but without toxic effects. Thus we assessed the antiviral activity of Baf-A1 at different concentrations (0.1-100 nM) in human alveolar epithelial cells (A549) infected with IAV strain A/PR/8/34 virus (H1N1). Infected and mock-infected cells pre- and cotreated with Baf-A1 were harvested 0-24 h postinfection and analyzed by immunoblotting, immunofluorescence, and confocal and electron microscopy. We found that Baf-A1 had disparate concentration-dependent effects on subcellular organelles and suppressed affected IAV replication. At concentrations ≥10 nM Baf-A1 inhibited acid lysosome formation, which resulted in greatly reduced IAV replication and release. Notably, at a very low concentration of 0.1 nM that is insufficient to reduce lysosome number, Baf-A1 retained the capacity to significantly impair IAV nuclear accumulation as well as IAV replication and release. In contrast to the effects of high concentrations of Baf-A1, very low concentrations did not exhibit cytotoxic effects or induce apoptotic cell death, based on morphological and FACS analyses. In conclusion, our results reveal that low-concentration Baf-A1 is an effective inhibitor of IAV replication, without impacting host cell viability.
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