Phosphoinositides (PIPnS) are integral signaling molecules for cellular communication. By recruiting specific proteins to the inner membrane, signal cascades are then initiated leading to downstream effects such as protein sorting, tumorigenesis, regulation of exo-and endocytosis, control of cell shape, cell migration and proliferation, and apoptosis. Discovering new PIPn specific lipid recognition proteins (LRPs) has become important in yielding new potential targets for drug discovery. Biochemical investigations on lipid selectivity have begun to employ protein overlay lipid blots, colloquially termed """"""""Fat Westerns"""""""", with nitrocellulose-immobilized PIPns. The converse approach would employ a lipid overlay with gel-separated proteins. Thus, in Phase I, we employed a liposome overlay system for identifying PIPn binding proteins immobilized on a nitrocellulose membrane using traditional PIPn-containing liposomes (PIPosomesTM). In addition, we developed a set of novel PIPn-containing polymerized liposomes named PolyPIPosomes TM. For several of the PIPn headgroups, we also investigated the use of PolyPIPosomes TM for detecting lipid-protein binding interactions. In Phase II, we propose to extend the methodology to include all eight PIPn scaffolds and perform stability studies. In addition we will further explore applications for PolyPIPosomes TM where regular liposomes have previously been used in basic research applications and in high throughput screening. Finally we will develop PolyPIPosomes TM and PIPosomes TM research products and establish collaborations for beta-testing products to facilitate their introduction to the market.
| Elkin, Sheryl K; Ivanov, Dmitri; Ewalt, Mark et al. (2005) A PHD finger motif in the C terminus of RAG2 modulates recombination activity. J Biol Chem 280:28701-10 |
