Cytosolic chloride ion is a key factor in lysosomal acidification and function of autophagy in human gastric cancer cell

The purpose of the present study was to clarify roles of cytosolic chloride ion (Cl(−)) in regulation of lysosomal acidification [intra-lysosomal pH (pH(lys))] and autophagy function in human gastric cancer cell line (MKN28). The MKN28 cells cultured under a low Cl(−) condition elevated pH(lys) and reduced the intra-lysosomal Cl(−) concentration ([Cl(−)](lys)) via reduction of cytosolic Cl(−) concentration ([Cl(−)](c)), showing abnormal accumulation of LC3II and p62 participating in autophagy function (dysfunction of autophagy) accompanied by inhibition of cell proliferation via G(0)/G(1) arrest without induction of apoptosis. We also studied effects of direct modification of H(+) transport on lysosomal acidification and autophagy. Application of bafilomycin A1 (an inhibitor of V-type H(+)-ATPase) or ethyl isopropyl amiloride [EIPA; an inhibitor of Na(+)/H(+) exchanger (NHE)] elevated pH(lys) and decreased [Cl(−)](lys) associated with inhibition of cell proliferation via induction of G(0)/G(1) arrest similar to the culture under a low Cl(−) condition. However, unlike low Cl(−) condition, application of the compound, bafilomycin A1 or EIPA, induced apoptosis associated with increases in caspase 3 and 9 without large reduction in [Cl(−)](c) compared with low Cl(−) condition. These observations suggest that the lowered [Cl(−)](c) primarily causes dysfunction of autophagy without apoptosis via dysfunction of lysosome induced by disturbance of intra-lysosomal acidification. This is the first study showing that cytosolic Cl(−) is a key factor of lysosome acidification and autophagy.