REGULATION OF THE PLASMA-MEMBRANE TYPE-III PHOSPHATIDYLINOSITOL 4-KINASE BY POSITIVELY CHARGED COMPOUNDS

The effects of positively charged compounds on a plasma membrane, type m phosphatidylinositol 4-kinase were studied. To determine whether th e enzyme would respond differently to the compounds in a membrane-asso ciated versus a soluble state, both the plasma membrane and solubilize d (released by 0.01% (v/v) Triton X-100) PI 4-kinase were used. Spermi dine, spermine, polylysine, cardiotoxin, melittin, and histone stimula ted the solubilized PI 4-kinase but had little effect on or weakly sti mulated the membrane-associated PI 4-kinase. Polyarginine inhibited me mbrane-associated PI 4-kinase 75% and inhibited the solubilized PI 4-k inase 30%, indicating that charge alone was not sufficient for activat ion. Polyarginine also eliminated the activation of the solubilized PI 4-kinase by a PI 4-kinase activator protein, PIK-A49. Calmodulin, a c ommon calcium-binding protein, at micromolar levels strongly inhibited solubilized PI 4-kinase activity but did not inhibit membrane-associa ted PI 4-kinase activity. The inhibition of,the solubilized PI 4-kinas e by calmodulin was calcium independent. Calcium alone (1 mu M-0.1 mM) inhibited PI 4-kinase activity only slightly (<30%). The differential effects of the positively charged compounds on the solubilized and me mbrane-associated PI 4-kinase were not due to substrate availability b ecause both enzymes were assayed in the presence of excess PI (0.6 mM) and 0.3% (v/v) Triton X-100. The data suggest that positively charged compounds affected the enzyme activity not only by interacting with t he substrates or products of the reaction but also by interacting with the PI 4-kinase or regulatory components in the plasma membrane. (C) 1994 Academic Press, Inc.