DESCRIPTION (provided by applicant): Anopheles mosquitoes are the primary vectors of malaria, one of humankind's most deadly and costly diseases. Malaria control measures are under increasing threat as drug- and insecticide- resistance spreads. Recently developed genomic resources and genetic methods provide unprecedented opportunities to improve our understanding of mosquito biology and develop new strategies to combat malaria and other mosquito-borne diseases. Y chromosome remains unexplored in Anopheles mosquitoes despite its importance in male determination and other aspects of male biology. Since blood feeding and, consequently, the transmission of disease pathogens is the sole province to female mosquitoes, it is critical to release only males when implementing genetic approaches for reducing mosquito populations or for replacing competent vector populations with populations that are refractory to disease transmission. Overcoming technical bottlenecks, we have recently uncovered four Y genes in An. stephensi. Among these, Guy1 is the best candidate for the male-determining M factor. Multiple Guy1-expressing transgenic lines were generated, which produced 6520 transgenic males and 0 transgenic females over multiple generations. Our preliminary data also suggest that the observed 100% female lethality in the Guy1 transgenic An. stephensi may result from mis-regulation of dosage compensation. Importantly, three known master switches of sex-determination, sex-lethal, Fem/Masc, xo-lethal 1, also directly or indirectly regulate dosage compensation in Drosophila melanogaster, Bombyx mori, and Caenorhabditis elegans, respectively. Loss of function sex- lethal alleles and knockdown of the target of Masc cause female embryonic lethality in D. melanogaster and B. mori, respectively. Here, we will test the hypothesis that the An. stephensi Guy1 is a master regulator of either sex-determination or dosage compensation or both. We will pursue the following specific aims: 1) To determine whether Guy1 controls dosage compensation and/or sex-determination; 2) To delineate the molecular mechanism underlying Guy1 function; and 3) To exploit Guy1 transgenics as a means for sex separation and population reduction in Anopheles mosquitoes.