Identification of common and cell type specific LXXLL motif EcR cofactors using a bioinformatics refined candidate RNAi screen in Drosophila melanogaster cell lines

The bioinformatics pre-screen of the Drosophila melanogaster proteome was successful in identifying an enriched putative candidate gene cohort. Over 80% of the genes tested yielded a positive hit in our reporter screen. We have identified both cell type specific and common cofactors which appear to be necessary for proper ecdysone induced gene regulation. We have determined that certain cofactors act as co-repressors to reduce target gene expression, while others act as co-activators to increase target gene expression. Interestingly, we find that a few of the cofactors shared among cell types have a reversible roles to function as co-repressors in certain cell types while in other cell types they serve as co-activators. Lastly, these proteins are highly conserved, with higher order organism homologs also harboring the LXXLL steroid receptor interaction domains, suggesting a highly conserved mode of steroid cell target specificity.In conclusion, we submit these cofactors as novel components of the ecdysone signaling pathway in order to further elucidate the dynamics of steroid specificity.Steroid hormones regulate many developmental processes in higher organisms, including postembryonic development, metamorphosis, and reproduction [1-3]. Pulses of the steroid hormone 20-hydroxyecdysone (referred to from here on as ecdysone) direct the morphological transitions of Drosophila throughout its life cycle [4-12]. Titers of ecdysone increase before each postembryonic larval molt and it is required for triggering metamorphosis transitions [3]. One of the highest peaks of ecdysone triggers the transition from third larval instar to puparium formation at the onset of metamorphosis, which involves simultaneous down-regulation of cell death inhibitors and up-regulation of cell death activators in larval tissues while activating proliferation and differentiation cascades in imaginal tissues [2,3,13]. This increase in the ecdysone titer during puparium formation is transduced to t