Research objectives and content
The discovery of inositol lipids, and the enzymes required to synthesize them, in the nucleus has led to the demonstration that the nuclear PI-cycle is regulated in a manner distinct from that at the plasma membrane. PtdIns(4P)5-kinase (Pipkinase) synthesizes Ptdlns(4,5)P2, a putative second messenger and a substrate-for three further second messenger molecules. Two distinct isoforms of this enzyme have been found in rat liver nuclei. This project proposes the characterization of both nuclear Pipkinase isoforms with a view to investigating the -regulation of these enzymes during the cell cycle. One isoforms is immunologically related to cytosolic Pipkinase C and monoclonal antibodies are available for immunopurification. The second isoform is immunologically unrelated and will be purified to homogeneity by column chromatography. Production of-monoclonal nuclear-specific antibodies will facilitate the cloning of both isoforms. Using these reagents, regulation of the nuclear Pipkinases will be studied during differentiation and cell cycle progression.
Training content (objective, benefit and expected impact)
The project provides training in eukaryotic cell biology techniques, lipid biochemistry and cellular signalling processes. With a strong grounding in prokaryotic systems and molecular biology I believe that experience in such a broad range of skills is integral to the development of a scientific career.