The Drosophila Toll pathway is involved in embryonic development, innate immunity, and cell-cell interactions. However, compared to the mammalian Toll-like receptor innate immune pathway, its intracellular signaling mechanisms are not fully understood. We have previously performed a series of ex vivo genome-wide RNAi screenings to identify genes required for the activation of the Toll pathway. In this study, we have conducted an additional genome-wide RNAi screening using the overexpression of Tube, an adapter molecule in the Toll pathway, and have performed a co-immunoprecipitation assay to identify components present in the dMyd88-Tube complex. Based on the results of these assays, we have performed a bioinformatic analysis, and describe candidate molecules and post-translational modifications that could be involved in Drosophila Toll signaling. (C) 2018 Elsevier Inc. All rights reserved.