Fig. 1
From: Targeting of VPS18 by the lysosomotropic agent RDN reverses TFE3-mediated drug resistance
a GO analysis of organelle-related genes according to cellular component. b The nuclear (Nuc) fractions of TFEB-expressing and TFE3-expressing PC3 cells treated with Doc (12 nM). Doxorubicin (Dox, 0.8 μM) was used as a positive control. c The Doc concentration in cytoplasm (Cyto), nuclear (Nuc), and lysosome (Lyso) was measured by high-performance liquid chromatography (HPLC). d Distribution of MRP2 and its colocalization with LAMP2 in PC3/Doc cells. e The expression of MRP2 and typical markers were detected by western blotting in TFE3-overexpressing cells. f Changes in the distribution of Doc in different organelles of TFE3-overexpressing cells. g Cells overexpressing TFE3 were treated with a combination of Doc (12 nM) and Dox (0.8 μM), and the expression of ABCC2 was analyzed by qPCR. h The synthesis of RDN-biotin, a chemical probe of RDN. i RDN-Bio (1 μM) interactors purified by streptavidin affinity purification were separted by SDS-PAGE. j Western blotting showing the coimmunoprecipitation of RDN-Bio (1 μM) with VPS18 but not with VPS16 and Biotin. k The interaction between VPS18 and VPS16 was disrupted by RDN-Bio (1 μM). l Model of human VPS18 mutants lacking three domains in the C-terminus (VPSΔ1), one domain in the C-terminus (VPSΔ2), two domains in the C-terminus (VPSΔ3), the RING domain (VPSΔ4), or the RING domain and the CC domain (VPSΔ5). m Western blotting showing the coimmunoprecipitation of RDN-Bio with overexpressed Flag-tagged full-length RDN, or the RING domain (VPSΔ4) or the RING with the CC domain (VPSΔ5) in 293T cells. n The expression of VPS18 in lung cancer (LUAD) and adjacent noncancerous tissues was detected by immunohistochemistry. o VPS18 is correlated with poor prognosis in human lung cancer. p Photographs of excised tumors from two groups (PC3/Doc-shNC and PC3/Doc-shVPS18-2) are shown. q Tumor volumes in different groups were recorded every 3 days. r The working model. TFE3/lysosome activation induced by therapeutics such as docetaxel (Doc) contributes to MDR by promoting lysosomal localization of MRP2 and enhancing drug sequestration in lysosomes. Lysosomal VPS18 as a potential target for the reversal of chemoresistance by a novel lysosomotropic agent RDN. Data are the mean ± SD; *p < 0.05, **p < 0.01, and ***p < 0.001
