LYSOSOMOTROPIC AGENTS ACTIVATE THE CAPACITY FOR CALCIUM DEPENDENT PINOCYTOSIS IN STARVED AMOEBA PROTEUS - EVIDENCE FOR A MECHANISM INVOLVING PHOSPHOLIPASE A(2)

摘要

Pinocytosis induced by Na+ was assayed by phase contrast microscopy in 8-12 days starved Amoeba proteus. These cultures were inactive with respect to calcium-dependent Na+-induced pinocytosis, but treatment with amino acid methyl and ethyl esters increased their capacity for pinocytosis. Besides promoting pinocytosis these compounds also stimulated calcium-sensitive secretion of lysosomal enzymes from normal, 2-3 days starved, cells. Only uncharged 1-forms of the amino acid esters were effective. Also other lysosomotropic compounds including monodansylcadaverine, glycine-phenylalanine-2-naphthylamide, NH4Cl, and the ionophores monensin and A23187 activated starved cells. The effect of these agents (except A23187) was inhibited by the drug dantrolene suggesting that activation is a consequence of release of Ca2+ from intracellular cellular stores. Several of the lysosomotropic agents also lost their activating effect in the presence of phospholipase A(2) (PLA(2)) inhibitors. To investigate whether or not PLA(2) activity in the cell culture could imitate the effect of the lysosomotropic agents, we incubated starved cells with snake venom PLA(2)s. These enzymes caused rapid, dantrolene-sensitive activation of the cells. Measurement of endogenous PLA(2) in ''normal'' cells revealed significant cellular activity but no significant secretion of the enzyme into the culture medium was observed. Together the studies with enzyme inhibitors and dantrolene suggest that the process by which lysosomotropic agents affect pinocytosis involves activation of PLA(2) and release of Ca2+ from intracellular stores. [References: 32]