The goal of this research is to understand the function, regulation, and localization of phosphatidylinositol 4-kinase (PI4K) in yeast (yeast Pik1).PI4K catalyzes the first committed step in the biosynthesis of PI4,4-bisphosphate (PI4,5P2) and, thus, is likely to be highly regulated. Hydrolysis of PI4,5P2 by Phospholipase C in response to extracellular stimuli generates two second messengers, inositol 1,4,5- trisphosphate and diacylglycerol (DAG), which initiates intracellular signaling cascades that modulate cell proliferation and differentiation. A number of PIK-related kinases have recently been identified and their functions are being unveiled. The dysfunction of PIKs leads to physiological disorders such as loss of immunological function and cancer. Revealing the function of these PIKs will contribute to our understanding and the cure of cancer. The PIK1 gene encoding a soluble PI4-kinase (125 Kd, 1066 aa) from S. cerevisiae was cloned and shown to be an essential gene in Thorner lab. The 274-residue C-terminal segment of Pik1 shares the highest homology with other PIKs including several PI3-kinases and comprise the catalytic domain. The 792-residue N- terminal segment of Pik1 contains, near its N-terminal end, a second region of high similarity between Pik1 and PI3-kinases. Evidence suggests that the activity of Pik1 is probably regulated through the interaction of its putative N-terminal regulatory domain with some other cellular protein(s). Rabbit polyclonal antisera against the N-terminal segment of Pik1 as well as an epitope-tagged version of this domain have been made. Two-hybrid assay and co-immunoprecipitation are being used to search for proteins that interact with Pik1 and to characterize their interaction. The localization of Pik1 is being investigated using both subcellular fractionation and indirect immunofluorescence. The minimal functional domains of Pik1 will be determined by deletion mutagenesis.
| Hendricks, K B; Wang, B Q; Schnieders, E A et al. (1999) Yeast homologue of neuronal frequenin is a regulator of phosphatidylinositol-4-OH kinase. Nat Cell Biol 1:234-41 |
