9304556 Boss The polyphosphorylated inositol phospholipids, phosphatidylinositol monophosphate (PIP) are present in the plasma membranes of higher plant cells; however, their functions in signal transduction are not well understood. PIP and PIP2 levels are relatively low in higher plants. This laboratory has isolated a 49 kDa protein from carrot cells that activates phosphatidylinositol (PI) 4-kinase, the enzyme that synthesizes PIP. The 49 kDa PI kinase activator, designated PIK-A49, is a multifunctional protein. It binds and bundles actin and has translational elongation factor-1-alpha activity. This research is based on the working hypothesis that PIK-A49 plays an important role in coordinating the status of PIP metabolism, the actin cytoskeleton, and protein synthesis within the cell. PIK-A49 and PI kinase are present in the plasma membrane, actin cytoskeleton and soluble fractions of carrot cells. Changes in the membrane and cytoskeletal PI kinase activity have been observed within seconds of exposing carrot cells to external stimuli. Whether PIP and PIP2 serve as precursors of second messengers or as direct membrane effectors, understanding the mechanism of regulation of PIP and PIP2 biosynthesis is essential in order to understand their roles in signal transduction. The goals of this research are to understand the mechanism of regulation of PIP biosynthesis in response to environmental stimuli and to understand how these changes in PIP metabolism might affect cell physiology. For these studies, carrot suspension culture cells will be used as a model system and two stimuli, one that increases PI 4-kinase activity and one that decreases PI 4-kinase activity. Specific questions to be addressed are: What is the mechanism of activation of PI 4-kinase by PIK-A49? Is there a change in the distribution of PIK-A49, PI kinase or F-actin in response to stimuli, and if so, what causes the change in distribution? This work provides a different perspective of PIP metabolism--one focused on PIP synthesis. In addition, the fact that the PIK-A49 activates PI 4-kinase, binds and bundles actin, and has elongation factor activity adds a new dimension to the potential for cross-talk among the plasma membrane, cytoskeleton, and protein synthesis machinery within plant cells during responses to environmental stimuli. %%% Little is known about the mechanisms used by plants to sense environmental signals. This investigator has shown that a subset of very negatively charged lipids in the outer membranes of plant cells undergo chemical changes in response to environmental stimuli. The biosynthesis of these lipids is regulated by an activator protein that can also regulate the structure of the cytoskeleton and protein synthesis within the cell. This finding suggested that the activator protein may serve to communicate the changes in the plant's environment from the outer membrane to the rest of the cell. The goals of this research are to understand how biosynthesis of these lipids is regulated and to determine whether the activator protein acts as a sensor to communicate environmental signals to the interior of the plant cell.***
