CHARACTERIZATION OF STRUCTURAL AND FUNCTIONAL PHOSPHOINOSITIDE DOMAINS IN HUMAN ERYTHROCYTE-MEMBRANES

In the erythrocyte membrane, only a fraction (50-60%) of phosphatidyli nositol 4,5-bisphosphate (PIP2) and of phosphatidylinositol 4-phosphat e (PIP) is rapidly turned over by specific kinases and phosphatases an d accessible to hydrolysis by the polyphosphoinositide (PPI)-specific phospholipase C (PLC). To investigate whether the metabolic segregatio n of PPI resulted from preferential interactions with proteins, we hav e measured the accessibility of PPI to bee venom phospholipase A2 (PLA 2) in native erythrocyte membranes, or after treatments designed to re move peripheral proteins and cytoplasmic domains of integral proteins. In native membranes, PPI, as well as the other major phospholipids, b ehaved as two distinct fractions (R1 and R2) differing by their sensit ivity to PLA2. Such a behavior was not observed in PIP and PIP2 contai ning artificial vesicles. Evidence was provided that the highly sensit ive fraction of PIP and PIP2 (R1) may be identical to the PLC-sensitiv e and rapidly metabolized pool. Removal of peripheral proteins, follow ed by proteolysis of the cytoplasmic domain of integral proteins, main ly glycophorins and band 3, led to a reduction of the R1 fraction of P IP and of PIP2. It is proposed that the rapidly metabolized pool Of PI P2 and PIP, involved in the regulation of major cellular functions, wo uld be maintained in its functional state through interactions with in tegral proteins.