Biochemical analysis of Torso and D-Raf during Drosophila embryogenesis: implications for terminal signal transduction

Determination of anterior and posterior terminal structures ofDrosophila embryos requires activation of two genes encoding putative protein kinases, torso and D-raf. In this study, we demonstrate that Torso has intrinsic tyrosine kinase activity and show that it is transiently tyrosine phosphorylated (activated) at syncytial blastoderm stages. Torso proteins causing a gain-of-function phenotype are constitutively tyrosine phospho-rylated, while Torso proteins causing a loss-of-function phenotype lack tyrosine kinase activity. The D-rafgene product, which is required for Torso function, is identified as a 90-kDa protein with intrinsic serine/threonine kinase activity. D-Raf is expressed throughout embryogenesis; however, the phosphorylation state of the protein changes during development. In wild-type embryos, D-Raf is hyperphosphorylated at 1 to 2 h after egg laying, and thereafter only the most highly phosphorylated form is detected. Embryos lacking Torso activity, however, show significant reductions in D-Raf protein expression rather than major alterations in the protein's phosphorylation state. This report provides the first biochemical analysis of the terminal signal transduction pathway in Drosophila embryos. In many multicellular organisms, receptor tyrosine ki-nases (RTKs) provide a means for cells to propagate and amplify signals received from the environment to specifi