Gene Editing Reveals Obligate and Modulatory Components of the CO2Receptor Complex in the Malaria Vector Mosquito,Anopheles coluzzii

The sensitivity to volatile carbon dioxide (CO 2 ) produced by humans and other animals is a critical component in the host preference behaviors of the malaria vector mosquito Anopheles coluzzii . The molecular receptors responsible for the ability to sense CO 2 are encoded by three putative gustatory receptor ( Gr ) genes ( Gr22,23,24 ) which are expressed in a distinctive array of sensory neurons housed in maxillary palp capitate peg sensilla of An. coluzzii . Despite the identification of these components and subsequent studies, there is a paucity of understanding regarding the respective roles of these three GRs in the mosquito’s CO 2 transduction process. To address this, we have used CRISPR/Cas9-based gene editing techniques combined with in vivo electrophysiological recordings to directly examine the role of Gr22,23,24 in detecting CO 2 in An. coluzzii . These studies reveal that both Gr23 and Gr24 are absolutely required to maintain in vivo CO 2 sensitivity while, in contrast, Gr22 knock out mutants are still able to respond to CO 2 stimuli albeit with significantly weaker sensitivity. Our data supports a model in which Gr22 plays a modulatory role to enhance the functionality of Gr23/24 complexes that are responsible for CO 2 sensitivity of mosquitoes.