Fig. 1: DELE1 negatively regulates mitophagy under mild mitochondrial stress conditions.
a Schematic illustration for monitoring mitophagy using mtKeima reporter via flow cytometry. Cells with increased 561 nm/405 nm mtKeima ratios compared to non-stressed cells were considered as mitophagy-positive. Created in BioRender. Guo, X. (https://BioRender.com/jqwlsdj). b PRKN expression levels were measured using flow cytometry as indicated by miRFP intensity. c HEK293T mtKeima reporter cells without overexpression (OE) of PRKN, d with a lower level (PRKN OELL), e with a medium level (PRKN OEML), f with a high level (PRKN OEHL), expressing a non-targeting control sgRNA (NTC) or sgRNA targeting DELE1 (DELE1 KD) were left untreated or treated with 5 or 10 μM CCCP, 0.6 or 1.25 ng/ml oligomycin, followed by measurement of mitophagy using flow cytometry. c–f show mean ± s.d., n = 3 independently treated culture wells, Two-way ANOVA test followed by Turkey’s multiple comparisons test (two-sided). **** adjusted p value < 0.0001. g PRKN OHHL with or without DELE1 KD were treated with 10 μM CCCP or 1.25 ng/ml oligomycin, followed by mitophagy measurement at 6 h and 24 h timepoints. (mean ± s.d., n = 3 independently treated culture wells) h Immunoblots of ATF4, LC3 and COX IV. PRKN OEHL cells with or without DELE1 KD were treated with 10 μM CCCP or 1.25 ng/mL of oligomycin for 24 h. β-actin was used as the loading control. i PRKN OELL and j PRKN OHHL cells with or without DELE1 KD were treated with antimycin A (0.4 and 2 μM) or rotenone (50 and 100 nM) for 24 hr, followed by measurement of mitophagy using flow cytometry. (mean ± s.d., n = 3 independently treated culture wells). Source data are provided as a Source Data file.
