EREZYILMAZ, Deniz F*; RIDDIFORD, Lynn M; TRUMAN, James W; University of Washington: An ancestral role for the metamorphosis-determining gene, broad in the direct-developing milkweed bug.

Fossil and phylogenetic evidence show that the metamorphosing insects diverged from direct-developing ancestors about 300 million years ago. To better understand the genetic basis of this divergence, we have focused on the role of the metamorphosis-determining gene, broad (br). In metamorphosing flies and moths, br expression is restricted to pupal development, and its expression at this time is required for production of the pupal stage. Genetic overexpression studies in Drosophila show that br acts by promoting the expression of pupal-specific genes, while suppressing the expression of larval- and adult-specific genes. We have cloned a homolog of this gene from the direct-developing milkweed bug, Oncopeltus fasciatus and have found that unlike its expression in flies and moths, Ofbr is expressed at each of the nymphal stages, but is absent at the molt to the adult stage. We used RNAi to test the role of nymphal Ofbr expression and found that Ofbr knockdown during nymphal development prevents heteromorphosis (the progression in pigment pattern that occurs between nymphal stages). Injected nymphs show normal growth, but simply repeat the pigment pattern at the stage of injection with Ofbr dsRNA. In addition, Ofbr is required for differential growth, as knockdown of Ofbr expression prevents anisometric growth of the wing pads, shifting the growth pattern of these structures into a more isometric mode, like that of the legs and antennae. Our data show that the ancestral role of br is to regulate deviations from simple isometric repetition during postembryonic growth. Our data further suggest that the role of br in regulating metamorphosis evolved by 1) restricting expression of this gene to one postembryonic instar, and 2) expansion of the number of traits that are regulated by br. This work was supported by NSF and NIH grants to JWT and LMR.