DEPENDENCE ON INTRACELLULAR PH AND MG2+ OF METABOLIC COMPARTMENTATIONOF PHOSPHOINOSITIDES IN HUMAN ERYTHROCYTES

Effects of intracellular pH and Mg2+ on turnover and extent of metabol ic compartmentation of phosphomonoester groups of phosphoinositides an d phosphatidate were investigated in human erythrocytes by short-term and equilibrium labeling with [P-32]P-i under steady-state conditions. At pH 6.7, the specific radioactivities of phosphoinositides reached apparent equilibrium values, in the range of 70% of that of ATP-gamma- P after long-term labeling. At pH 7.2, these values were in the range of 40-50% of that of ATP-gamma-P, This demonstrates a decreased access ibility of phosphoinositides to enzymatic phosphorylation and dephosph orylation at the transition from acidic to normal incubation condition s. These changes were more pronounced at pH 7.8. High intracellular [M g2+] initially activated the turnover of phosphoinositides at pH 7.2. The activation changed into an inhibition and an increase of metabolic compartmentation after three hours preincubation of erythrocytes at h igh [Mg2+]. The long-term Mg2+ effects are reversible to a great exten t by a subsequent re-reduction of [Mg2+]. In conclusion, deprotonation of phosphoinositides by low [H+] or high intracellular [Mg2+] at norm al pH induces a decreased accessibility for their specific lipid kinas es and phosphatases. This effect may be the result of lateral phase se paration of acidic phospholipids as a consequence of divalent cation c omplexation under both experimental conditions, high pH and high [Mg2] at normal pH.