Functional analysis of the ABCs of eye color in Helicoverpa armigera with CRISPR/Cas9-induced mutations.

Many insect pigments are localized in subcellular pigment granules, and transport of pigment precursors from the cytoplasm is accomplished by ABC proteins. Drosophila melanogaster has three half-transporter genes (white, scarlet, and brown, all affecting eye pigments) and Bombyx mori has a fourth (ok). The White, Brown, Scarlet and Ok proteins each have one transmembrane and one cytoplasmic domain and they heterodimerize to form functional transporters with different substrate specificities. We used CRISPR/Cas9 to create somatic and germ-line knockout mutations of these four genes in the noctuid moth Helicoverpa armigera. Somatic knockouts of white block pigmentation of the egg, first instar larva and adult eye, but germ-line knockouts of white are recessive lethal in the embryo. Knockouts of scarlet are viable and produce pigmentless first instar larvae and yellow adult eyes lacking xanthommatin. Knockouts of brown show no phenotypic effects on viability or pigmentation. Knockouts of ok are viable and produce translucent larval cuticle and black eyes. CRISPR/Cas9-induced mutations are a useful tool for analyzing how essential and non-essential genes interact to produce the diversity of insect pigmentation patterns found in nature.