DISINTEGRATION OF LYSOSOMES MEDIATED BY GTP-GAMMA-S-TREATED CYTOSOL -POSSIBLE INVOLVEMENT OF PHOSPHOLIPASES

We showed previously that cytosol treated with guanosine 5'-O-(3-thiot riphosphate) (GTP gamma S) disintegrated lysosomes in vitro [Sai, Y. e t al. (1994) Biochem. Biophys. Res. Commun. 198, 869-877] in time-, te mperature-, and dose-dependent manners. This also requires ATP, howeve r, the latter can be substituted with deoxy-ATP, ADP, or ATP gamma S, suggesting no requirement of ATP hydrolysis. The lysis was inhibited b y several chemical modifiers, including N-ethylmaleimide, 7-chloro-4-n itrobenzo-2-oxa-1,3-diazole, and 4,4'-diisothio-cyanatostilbene-2,2'-d isulfonic acid, and by various phospholipase inhibitors (trifluoperazi ne, p-bromophenacyl bromide, nordihydroguaiaretic acid, W-7, primaquin e, compound 48/80, neomycin, and gentamicin), but not by ONO-RS-082, a n inhibitor of phospholipase A:. The reaction was also inhibited by ph ospholipids (phosphatidylinositol, phosphatidylserine, phosphatidic ac id, and phosphatidylcholine) and diacylglycerol. Among the phospholipa se A(2) hydrolysis products Of phospholipids, unsaturated fatty acids (oleate, linoleate, and arachidonate) and lysophospholipid (lysophosph atidylcholine) by themselves broke lysosomes down directly, whereas sa turated fatty acids (palmitate and stearate) had little effect. We fou nd that GTP gamma S-stimulated cytosolic phospholipase A(2) activity w as highly sensitive to ONO-RS-082. These results suggest the participa tion of phospholipase(s), though not cytosolic phospholipase A(2), in the GTP gamma S-dependent lysis of lysosomes.