Sex-specific trait expression represents a striking dimension of morphological variation within and across species. The mechanisms instructing sex-specific organ development have been well studied in a small number of insect model systems, suggesting striking conservation in some parts of the somatic sex determination pathway while hinting at possible evolutionary lability in others. However, further resolution of this phenomenon necessitates additional taxon sampling, particularly in groups in which sexual dimorphisms have undergone significant elaboration and diversification. Here, we functionally investigate the somatic sex determination pathway in the gazelle dung beetle Digitonthophagus gazella, an emerging model system in the study of the development and evolution of sexual dimorphisms. We find that RNA interference (RNAi) targeting transformer (tra) caused chromosomal females to develop morphological traits largely indistinguishable from those normally only observed in males, and that tra is sufficient to induce splicing of the normally male-specific isoform of doublesex in chromosomal females, while leaving males unaffected. Further, intersex was found to phenocopy previously described RNAi phenotypes of doublesex in female but not male beetles. These findings match predictions derived from models of the sex determination cascade as developed largely through studies in Drosophila melanogaster. In contrast, efforts to target transformer2 via RNAi resulted in high juvenile mortality but did not appear to affect doublesex splicing, whereas RNAi targeting Sex-lethal and two putative orthologs of hermaphrodite yielded no obvious phenotypic modifications in either males or females, raising the possibility that the function of a subset of sex determination genes may be derived in select Diptera and thus nonrepresentative of their roles in other holometabolous orders. Our results help illuminate how the differential evolutionary lability of the somatic sex determination pathway has contributed to the extraordinary morphological diversification of sex-specific trait expression found in nature.
