Regulated assembly of the Toll signaling complex drives Drosophila dorsoventral patterning

In Drosophila, the Toll pathway establishes the embryonic dorsoventral axis and triggers innate immune responses to infection. The transmembrane receptor Toll acts through three death domain-containing proteins, the kinase Pelle and the adapters Tube and MyD88, in signaling to downstream NF-κB-like transcription factors. Here, we delineate the critical events in the earliest stages of Toll signaling. Mutational studies based on structural modeling reveal that the direct interaction of the bivalent Tube death domain with MyD88 is critical for signaling in vivo. The complex of MyD88 and Tube forms prior to signaling and is localized to the embryonic plasma membrane by MyD88. Upon Toll homodimerization, this complex is rapidly recruited to Toll. Binding of Pelle to the MyD88–Tube complex promotes Pelle activation, leading to degradation of the IκB-like inhibitor, Cactus. Together, these experiments convert a linear picture of gene function into a dynamic mechanistic and structural understanding of signaling complex assembly and function

A Gradient of Cactus Protein Degradation Establishes Dorsoventral Polarity in theDrosophilaEmbryo

AbstractDorsoventral polarity in theDrosophilaembryo is established by a signaling pathway active on the ventral and ventrolateral surfaces of the embryo. Signal transduction via the protein kinase Pelle frees the Rel-related protein Dorsal from its cytoplasmic inhibitor Cactus, allowing Dorsal to translocate into ventral and ventrolateral nuclei and direct gene expression. Here, we show by immunochemical analyses that Pelle-mediated signaling induces the spatially graded degradation of Cactus. Using a tissue culture system which reconstitutes Pelle-dependent Cactus degradation, we show that a motif in Cactus resembling the sites of signal-dependent phosphorylation in the vertebrate homologs IκB-α and IκB-β is essential for Pelle-induced Cactus degradation. Substitution of four serines within this motif with nonphosphorylatable alanine residues generated a mutant Cactus that still functions as a Dorsal inhibitor but is resistant to induced degradation. Injection of RNA encoding this altered form of Cactus has a dominant negative effect on establishment of dorsoventral polarity in the embryo. We conclude that dorsoventral signaling results in a Cactus concentration gradient and propose that signal-dependent phosphorylation directs the spatially regulated proteolysis of Cactus protein