Phosphorylation modulates direct interactions between the Toll receptor, Pelle kinase and Tube

Determination of dorsal/ventral polarity in Drosophila requires 12 genetica lly defined, maternally encoded proteins. These include Toll, a transmembra ne receptor, Pelle, a ser/thr protein kinase and Tube, all of which functio n intracytoplasmically to initiate the cascade that ultimately activates Do rsal, an NF-kappa B family transcription factor. Here we describe biochemic al interactions between recombinant Toll, Pelle and Tube that provide insig hts into early events in activation of the signaling cascade. We first show that Pelle binds directly to a region within the Toll intracytoplasmic dom ain, providing the first evidence that these two evolutionarily conserved m olecules physically interact. We then demonstrate that Pelle can be autopha sphorylated, and that this prevents binding to Toll as well as Tube. Autoph osphorylation occurs in the N-terminal, death-domain-containing region of P elle, which is dispensable for binding to Toll but required for enzymatic a ctivity. We also show that Pelle phosphorylates Toll, within the region req uired for Pelle interaction, but this phosphorylation can be blocked by a p reviously characterized inhibitory domain at the Toll C terminus. These and other results allow us to propose a model by which multiple phosphorylatio n-regulated interactions between these three proteins lead to activation of the Dorsal signaling pathway.