APEX1 is identified as a novel Mode I TFEB regulator. (A) Quantification of TFEB::mNG nuclear-translocated cells in WT HeLa cells stably expressing TFEB::mNG, transfected with scrambled siRNA (siCon) or the indicated siRNA. The cells were treated with 1 mM LLOMe for 1 h (n = 3 biologically independent samples). (B) Representative immunoblots of TFEB, p-TFEB (Ser211), APEX1, and GAPDH in WT HeLa cells under the indicated conditions. Cells were transfected with siLuc and siAPEX1. (C) Quantification of image data of p-TFEB (Ser211)/TFEB shown in B (n = 3 biologically independent samples). (D) Results of the PLA using ATG7 KO HeLa cells stably expressing TFEB::EGFP. The indicated plasmids were transiently transfected and were either untreated (control) or treated with 1 mM LLOMe for 1 h followed by the PLA. (E) Representative fluorescence images of WT HeLa cells transiently expressing TFE3::EGFP or MITF::EGFP, either untreated (control) or treated with 1 mM LLOMe for 3 h. (F) Quantification of image data shown in E (n = 3 biologically independent samples). The relative values of nuclear/cytoplasmic ratios of TFE3 or MITF::EGFP compared with the control are shown. (G) Results of the PLA in WT HeLa cells expressing TFE3::EGFP or MITF::EGFP. The indicated plasmids were transiently transfected and were either untreated (control) or treated with 1 mM LLOMe for 3 h followed by the PLA. (H) Representative fluorescence images of HeLa cells expressing mNG::APEX1 (green) and either untreated (control) or treated with 1 mM LLOMe for 1 h. (D, E, G, and H) Scale bars, 50 µm. (E) Arrowheads indicate nuclear-localized TFE3::EGFP or MITF::EGFP. (C and F) P values were determined by ANOVA with Tukey’s multiple comparison test (C) or t test (F); *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Source data are available for this figure: SourceData FS3.
Figure S3.

APEX1 is identified as a novel Mode I TFEB regulator. (A) Quantification of TFEB::mNG nuclear-translocated cells in WT HeLa cells stably expressing TFEB::mNG, transfected with scrambled siRNA (siCon) or the indicated siRNA. The cells were treated with 1 mM LLOMe for 1 h (n = 3 biologically independent samples). (B) Representative immunoblots of TFEB, p-TFEB (Ser211), APEX1, and GAPDH in WT HeLa cells under the indicated conditions. Cells were transfected with siLuc and siAPEX1. (C) Quantification of image data of p-TFEB (Ser211)/TFEB shown in B (n = 3 biologically independent samples). (D) Results of the PLA using ATG7 KO HeLa cells stably expressing TFEB::EGFP. The indicated plasmids were transiently transfected and were either untreated (control) or treated with 1 mM LLOMe for 1 h followed by the PLA. (E) Representative fluorescence images of WT HeLa cells transiently expressing TFE3::EGFP or MITF::EGFP, either untreated (control) or treated with 1 mM LLOMe for 3 h. (F) Quantification of image data shown in E (n = 3 biologically independent samples). The relative values of nuclear/cytoplasmic ratios of TFE3 or MITF::EGFP compared with the control are shown. (G) Results of the PLA in WT HeLa cells expressing TFE3::EGFP or MITF::EGFP. The indicated plasmids were transiently transfected and were either untreated (control) or treated with 1 mM LLOMe for 3 h followed by the PLA. (H) Representative fluorescence images of HeLa cells expressing mNG::APEX1 (green) and either untreated (control) or treated with 1 mM LLOMe for 1 h. (D, E, G, and H) Scale bars, 50 µm. (E) Arrowheads indicate nuclear-localized TFE3::EGFP or MITF::EGFP. (C and F) P values were determined by ANOVA with Tukey’s multiple comparison test (C) or t test (F); *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Source data are available for this figure: SourceData FS3.

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