| The Ecdysoneless (Ecd) protein is highly conserved from yeast to human, indicative of a conserved function through sequences retained through evolution. At present, however, no specific structural domains or motifs linked to function have been identified in the Ecd protein of any species. Preliminary analysis of loss-of-function phenotypes of Ecd across evolution has demonstrated a fundamental role of this protein in controlling cell growth/survival, and development in a cell-autonomous fashion. The important roles of Ecd in controlling basic biochemical pathways were further substantiated by our studies in mammals where Ecd knockout mice were lethal at early embryonic stage.;To understand the cellular function of mammalian Ecd, Ecd floxed/floxed embryo fibroblasts were used for functional studies of the Ecd gene using the Cre-mediated gene deletion strategy. Importantly, Ecd regulates cell cycle progression by modulating the Rb/E2F pathway. Furthermore, Ecd interacts with Rb family proteins and competes with E2F for interaction with Rb. Defective cell cycle progression in Ecd-null cells is accompanied by impaired Rb dissociation from E2F target gene promoters, and the down-regulation of E2F target gene expression, which are essential for cell cycle progression. Thus, this finding indicates that mammalian Ecd is essential for embryonic development in mice and plays a critical role in mammalian cell cycle regulation.;In order to explore other potential roles of mammalian Ecd, Ecd interacting proteins were identified by using tandem affinity purification followed by mass spectrometry. I identified several potential Ecd-interacting proteins that are involved in various cellular pathways such as transcription, mRNA splicing, ATP transport, and metabolism implicating Ecd in several important biological functions, which are currently under study in my mentor's laboratory. For example, my biochemical analysis using reporter assay proved that mammalian Ecd has an intrinsic transactivation activity. Furthermore, the co-operativity of Ecd with p300, a global transcriptional coactivator, in its transactivation activity suggests that Ecd could play a role as a transcriptional regulator. Lastly, the mutational analysis of the Ecd protein will be extremely helpful in further defining its transactivation function, subcellular localization and protein-protein interaction to fully define physiological functions of mammalian Ecd. |
