Investigation of Isoform Specific Functions of the V-ATPase a Subunit During Drosophila Wing Development

The vacuolar ATPases (V-ATPases) are ATP-dependent proton pumps that play vital roles in eukaryotic cells. Insect V-ATPases are required in nearly all epithelial tissues to regulate a multiplicity of processes including receptor-mediated endocytosis, protein degradation, fluid secretion, and neurotransmission. Composed of fourteen different subunits, several V-ATPase subunits exist in distinct isoforms to perform cell type specific functions. The 100 kD a subunit (Vha100) of V-ATPases are encoded by a family of five genes inDrosophila, but their assignments are not fully understood. Here we report an experimental survey of theVha100gene family duringDrosophilawing development. A combination of CRISPR-Cas9 mutagenesis, somatic clonal analysis andin vivoRNAi assays is used to characterize the requirement of Vha100 isoforms, and mutants ofVha100-2,Vha100-3, Vha100-4, andVha100-5genes were generated. We show thatVha100-3andVha100-5are dispensable for fly development, whileVha100-1is not critically required in the wing. As for the other two isoforms, we find thatVha100-2regulates wing cuticle maturation, whileVha100-4is the single isoform involved in developmental patterning. More specifically,Vha100-4is required for proper activation of the Wingless signaling pathway during fly wing development. Interestingly, we also find a specific genetic interaction betweenVha100-1andVha100-4during wing development. Our results revealed the distinct roles ofVha100isoforms during insect wing development, providing a rationale for understanding the diverse roles of V-ATPases.