Distinct activation patterns of EGF receptor signaling in the homoplastic evolution of eggshell morphology in genus Drosophila

Homoplasy is a phenomenon in which organisms in different phylogenetic groups independently acquire similar traits. However, it is largely unknown how developmental mechanisms are altered to give rise to homoplasy. In the genus Drosophila, all species of the subgenus Sophophora, including Drosophila (D.) melanogaster, have eggshells with two dorsal appendages (DAs); most species in the subgenus Drosophila, including D. virilis, and in the subgenus Dorsilopha, have four-DAs. D. melanica belongs to the Drosophila subgenus, but has two-DAs, and phylogenetic analyses suggest that it acquired this characteristic independently. The patterning of the DAs is tightly regulated by epidermal growth factor receptor (EGFR) signaling in D. melanogaster. Previous studies suggested that a change in the EGFR signal activation pattern could have led to the divergence in DA number between D. melanogaster and D. virilis. Here, we compared the patterns of EGFR signal activation across the Drosophila subgenera by immunostaining for anti-activated MAP kinase (MAPK). Our analysis revealed distinct patterns of EGFR signal activation in each subgenus that was consistent with their phylogenetic relationship. In addition, the number of DAs always corresponded to the number of EGFR signaling activation domains in two, three, and four-DA species. Despite their common two-DA characteristic, the EGFR signaling activation pattern in D. melanica diverged significantly from that of species in the subgenus Sophophora. Our results suggest that acquisition of the homoplastic two-DA characteristic could be explained by modifications of the EGFR signaling system in the genus Drosophila that occurred independently and at least twice during evolution.