Phosphoinositides and lipid kinases in oxidative stress signalling and cancer

Phosphoinositides are implicated in virtually all aspects of cellular welfare. They help to maintain the structure of biomembranes, but are also essential signal transduction molecules. This is illustrated by PtdIns(4,5)P2, a versatile and important phosphoinositide that regulates many cellular processes including cell survival, calcium homeostasis, vesicular trafficking, cytokinesis, stress responses and cell adhesion. Cells use various enzymes to regulate PtdIns(4,5)P2 levels, including lipid kinases, lipid phosphatases and lipases. PtdIns(4,5)P2 can be synthesized by two closely related families of lipid kinases. Phosphatidylinositol-4-phosphate (PtdIns4P) 5-kinase (PIP5K) phosphorylates PtdIns4P and phosphatidylinositol-5-phosphate (PtdIns5P) 4-kinase (PIP4K) phosphorylates PtdIns5P to make PtdIns(4,5)P2. Cellular PtdIns4P is approximately ten times more abundant than PtdIns5P and therefore PtdIns(4,5)P2 is likely generated by PIP5K mainly. In vivo studies in animals and in mammalian cells suggest that PIP4Ks primarily regulate the level of PtdIns5P, although they may also regulate a specific pool of PtdIns(4,5)P2. PtdIns5P is the latest addition to the ‘palet’ of phosphoinositides and it is the least understood one. In this thesis, I have examined biochemical and functional aspects of PtdIns5P and the PIP4Ks that determine its levels in the cell. I have investigated how PtdIns5P levels change in response to redox signaling, or in response to the activity of the isoforms PIP4K2A and PIP4K2B and I have evaluated PIP4K2B expression in breast tumours.