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

Abstract

The purpose of the present study was to clarify roles of cytosolic chloride ion (Cl−) in regulation of lysosomal acidification [intra-lysosomal pH (pHlys)] and autophagy function in human gastric cancer cell line (MKN28). The MKN28 cells cultured under a low Cl− condition elevated pHlys 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 G0/G1 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 pHlys and decreased [Cl−]lys associated with inhibition of cell proliferation via induction of G0/G1 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.

A synthetic ion transporter that disrupts autophagy and induces apoptosis by perturbing cellular chloride concentrations

“…Dysregulation of intracellular chloride concentrations can lead to suppression of cell proliferation via autophagy disruption by altering the lysosomal pH and impairing lysosomal enzyme activity 14,46 . Autophagy starts with engulfment of cytoplasmic constituents by the phagophore, which is then expanded to form the double-membrane autophagosome (Supplementary Fig.…”

“…It is known that the dysregulation of intracellular concentrations of ions, particularly chloride anions, correlates closely with the onset of apoptosis 13 . Physiological perturbations in cellular chloride concentrations can also affect pH and autophagy 14 . Lysosomal pH values are typically maintained at <5.0 by sustaining a ~80 mM lysosomal chloride concentration through the action of the lysosomal chloride transporters 11 .…”

Fluorouracil induces autophagy-related gastric carcinoma cell death through Beclin-1 upregulation by miR-30 suppression

“…Currently, surgical treatment is the main therapy of GC, although the outcome of the treatment is far from satisfaction [8][9][10][11]. Chemotherapy, especially fluorouracil (5-FU), is used as a supplementary treatment for GC and has resulted in better survival of the patients [12][13][14][15][16]. However, the molecular mechanisms underlying the anti-cancer effects of 5-FU are not completely understood.…”

A cyclohexadepsipeptide from entomogenous fungi Metarhizium anisopliae inhibits the Helicobacter pylori induced pathogenesis through attenuation of vacuolating cytotoxin-A activity

“…The molecular actions of Baf A1-induced apoptosis of cancer cells were included increasing lysosomal pH and activation of caspase-3 [33]. Additionally, a very recent study reported that treatment of gastric cancer with Baf A1, decreased the lysosomal chloride and therefore causes dysfunction of autophagy [34]. In the study of Dtx-B which harbored the same activity with Baf A1 that have been demonstrated bearing apoptosis effects on several types of cancers including lung cancer [18], colorectal cancer [35], and lymphoma [36].…”