Signalling and non-caveolar rafts

Rafts are small membrane domains containing discrete subsets of lipids and proteins. Although microscopic raft structures termed ‘caveolae’ were described nearly 50 years ago, the importance of rafts, particularly signalling within rafts, is only beginning to be understood. Our studies focus on receptor-dependent phosphoinositide signalling. Using their characteristic buoyancy in density gradients, we and others found that the epidermal growth factor (EGF) receptor, phosphatidyl-inositol 4-kinase and phosphoinositides are localized within a caveolin-rich fraction of A431 carcinoma cells. We subsequently found that membrane fragments containing the EGF receptor and most cellular phosphoinositides can be separated from caveolae. Consequently, components of EGF-dependent phosphoinositide signalling localize to one or more novel types of raft, the composition of which we are currently determining. A key component is the type II phosphatidylinositol 4-kinase, which, for many years, has proven difficult to purify and clone. We describe our recent purification from rafts and cloning of this elusive enzyme, and discuss how the structure sheds light on the rafting of this enzyme.