Functional analysis of JAK/STAT signalling using genome-wide RNAi

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The JAK/STAT signal transduction pathway is involved in development, immunity anddisease and has been conserved throughout evolution. While components of both theDrosophila and mammalian JAK/STAT pathway have been characterized by biochemical andgenetic means, a number of significant gaps in our understanding of the pathway remain. Herewe propose experiments to comprehensively identify the genes involved in poorly understoodaspects of pathway signalling using Drosophila as a model system. First, using an availableRNAi libary targeting every gene in the fly genome, we will sreen for JAK/STATcomponents in Drosophila macrophage-like cells. To this end, we have established a reportergene assays that accurately reflects JAK/STAT pathway activity. We will then re-screen thepotentially interacting genes identified during the initial screen to determine those directlyimplicated in our specific fields of research. These interacting genes will be evaluated andprioritized before analysis using genetic and biochemical experiments. Martin Zeidler willfurther identify genes involved in upstream processes of ligand modification, secretion,receptor binding and internalisation. This will employ a GFP-tagged ligand which can bevisualised both in cultured cells in vivo. Mutations in the genes affecting these processes willbe analysed to determine their interactions with JAK/STAT dependent phenotypes in vivo.Michael Boutros will analyze newly identified genes for their role during innate immunity.JAK/STAT signalling have been implicated by us as part of the Drosophila innate immuneresponse, although its in vivo function remains poorly understood. Loss-of-functionphenotypes of novel components will be studied in vitro.