The inhibition of chloride intracellular channel 1 enhances Ca and reactive oxygen species signaling in A549 human lung cancer cells.

Chloride intracellular channel 1 (CLIC1) is a promising therapeutic target in cancer due to its intrinsic characteristics; it is overexpressed in specific tumor types and its localization changes from cytosolic to surface membrane depending on activities and cell cycle progression. Ca and reactive oxygen species (ROS) are critical signaling molecules that modulate diverse cellular functions, including cell death. In this study, we investigated the function of CLIC1 in Ca and ROS signaling in A549 human lung cancer cells. Depletion of CLIC1 via shRNAs in A549 cells increased DNA double-strand breaks both under control conditions and under treatment with the putative anticancer agent chelerythrine, accompanied by a concomitant increase in the p-JNK level. CLIC1 knockdown greatly increased basal ROS levels, an effect prevented by BAPTA-AM, an intracellular calcium chelator. Intracellular Ca measurements clearly showed that CLIC1 knockdown significantly increased chelerythrine-induced Ca signaling as well as the basal Ca level in A549 cells compared to these levels in control cells. Suppression of extracellular Ca restored the basal Ca level in CLIC1-knockdown A549 cells relative to that in control cells, implying that CLIC1 regulates [Ca] through Ca entry across the plasma membrane. Consistent with this finding, the L-type Ca channel (LTCC) blocker nifedipine reduced the basal Ca level in CLIC1 knockdown cells to that in control cells. Taken together, our results demonstrate that CLIC1 knockdown induces an increase in the intracellular Ca level via LTCC, which then triggers excessive ROS production and consequent JNK activation. Thus, CLIC1 is a key regulator of Ca signaling in the control of cancer cell survival.