The gap junction protein Innexin3 is required for eye disc growth in Drosophila

The Drosophila compound eye develops from a bilayered epithelial sac composed of an upper peripodial epithelium layer and a lower disc proper, the latter giving rise to the eye itself. During larval stages, complex signalling events between the layers contribute to the control of cell proliferation and differentiation in the disc. Previous work in our lab established the gap junction protein Innexin2 (Inx2) as crucial for early larval eye disc growth. By analysing the contribution of other Innexins to eye size control, we have identified Innexin3 (Inx3) as an important growth regulator. Depleting inx3 during larval eye development reduces eye size, while elevating inx3 levels increases eye size, thus phenocopying the inx2 loss- and gain-of-function situation. As demonstrated previously for inx2, inx3 regulates disc cell proliferation and interacts genetically with the Dpp pathway, being required for the proper activation of the Dpp pathway transducer Mad at the furrow and the expression of Dpp receptor Punt in the eye disc. At the developmental timepoint corresponding to eye disc growth, Inx3 colocalises with Inx2 in disc proper and peripodial epithelium cell membranes. In addition, we show that Inx3 protein levels critically depend on inx2 throughout eye development and that inx3 modulates Inx2 protein levels in the larval eye disc. Rescue experiments demonstrate that Inx3 and Inx2 cooperate functionally to enable eye disc growth in Drosophila. Finally, we demonstrate that expression of Inx3 and Inx2 is not only needed in the disc proper but also in the peripodial epithelium to regulate growth of the eye disc. Our data provide a functional demonstration that putative Inx2/Inx3 heteromeric channels regulate organ size.