| Phosphoinositides play important roles in intracellular trafficking and signaling. Endocytosis of activated growth factor receptors is a major route for termination of receptor signaling and is critically dependent on phosphatidylinositol 4,5-bisphosphate (PIP2) and its downstream product phosphatidylinositol 3,4,5-trisphosphate (PIP3). Recent data suggest that at least some growth factor receptor signals can be transduced from endosomal compartments. Activated receptors are sorted at early endosomes to be targeted for recycling (signaling continuation) or lysosomal degradation (signal termination). Hence, endosomal sorting influences the duration of signaling, its amplitude, and possibly quality (i.e. by differential effector recruitment at plasmalemmal vs endosomal membranes) and thus has important implications for cell differentiation, cell cycle progression and cancer. Outstanding key questions addressed by the proposed research are (i) how PIP3 regulates growth factor trafficking at the plasmalemma and at endosomes, and (ii) which signaling pathways are elicited by cell surface vs endosomally produced PIP3. By adapting a recently developed technique to rapidly deplete specific pools of phosphoinositides at select membranes, I plan to deplete PIP3 at different intracellular (i.e. endosomal) locations to study its effect on growth factor-induced signaling cascades and receptor trafficking, using biochemical and state-of-the-art imaging approaches including spinning disc confocal and FRET microscopy. |
