Regulation of phosphatidylinositol 4-kinase by the protein activator PIK-A49. Activation requires phosphorylation of PIK-A49.
PIK-A49 is a 49-kDa soluble protein that was isolated as an activator of the plasma membrane phosphatidylinositol (PI) 4-kinase from carrot cells (Yang, W., Burkhart, W., Cavallius, J., Merrick, W. C., and Boss, W. F. (1993) J. Biol. Chem. 268, 392-398). PIK-A49 is a multifunctional protein that binds and bundles F-actin and has translational elongation factor-1 alpha activity. In this paper, we have investigated the mechanism of activation of PI 4-kinase by PIK-A49. PIK-A49 decreased the Km of PI 4-kinase for ATP from 0.40 to 0.19 mM. GTP and GDP, which affect the elongation factor-1 alpha function of the protein, inhibited the activation of PI 4-kinase by PIK-A49. Phosphorylation of purified PIK-A49 by a calcium-dependent protein kinase enhanced activation of PI 4-kinase. When dephosphorylated by alkaline phosphatase, PIK-A49 no longer activated PI 4-kinase; however, rephosphorylation of PIK-A49 by calcium-dependent protein kinase fully restored activation. Western blots using anti-PIK-A49 serum showed that PIK-A49 was associated with the plasma membrane and the F-actin fraction isolated from plasma membranes, indicating that PIK-A49 would be in a position to regulate plasma membrane PI 4-kinase. Based on these data, we propose a mechanism for feed-forward regulation of polyphosphoinositide biosynthesis in response to increases in cytosolic calcium.
Yang
W
W
Botany Department, North Carolina State University, Raleigh 27695-7612.
Boss
W F
WF
D019870:1-Phosphatidylinositol 4-Kinase; D002118:Calcium / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D004533:Egtazic Acid / Q000494:pharmacology; D004789:Enzyme Activation; D006153:Guanosine Diphosphate / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D007700:Kinetics; D008970:Molecular Weight; D010445:Peptide Elongation Factors / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000302:isolation & purification / Q000378:metabolism*; D010940:Plant Proteins / Q000302:isolation & purification / Q000378:metabolism*; D010944:Plants / Q000201:enzymology*; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism
C085842:PIK-A49 protein, Daucus carota; D010445:Peptide Elongation Factors; D010940:Plant Proteins; D006153:Guanosine Diphosphate; D004533:Egtazic Acid; D006160:Guanosine Triphosphate; D011494:Protein Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D019870:1-Phosphatidylinositol 4-Kinase; D002118:Calcium