Fig. 5: NRF1 induces mitophagy during OGD/R. | Cell Death Discovery

Fig. 5: NRF1 induces mitophagy during OGD/R.

From: NRF1 coordinates mitochondrial adaptations to dampen intracellular ROS and inflammatory responses during ischemia reperfusion

Fig. 5

A Heatmap indicating relative expression levels of proteasome subunit genes differentially expressed between WT and NFE2L1−/− BMDMs subject to OGD/R. B-C. WT and NFE2L1−/− BMDMs were subject to OGD/R followed by analysis of poly-ubiquitinated proteins by WB (B). Intensityof Ubiquitin was normalized to α-Tubulin (C). D, E WT and NFE2L1−/− BMDMs were subject to OGD/R followed by analysis of NRF1 in cytosolic and mitochondrial fractions by WB (D). Intensity of Ubiquitin was normalized to α-Tubulin in cytoplasm and VDAC1 in mitochondria (E). F, G WT and NFE2L1−/− BMDMs were subject to OGD/R +/− Bafilomycin A1 (1 nM) treatment during reoxygenation. LC3 lipidation in mitochondrial fractions was assessed by WB (F). Data are presented as fold induction of LC3-II band intensity in the +BafA1 condition versus the -BafA1 condition for the corresponding OGD/R stimulation time point (E). H, I WT and NFE2L1−/− BMDMs were subject to OGD/R +/− Bafilomycin A1 (1 nM) treatment during reoxygenation. LC3 puncta and mitochondria co-localization was assessed by immunofluorescent microscopy. Quantitation (H) and representative images (I) are shown. J, K WT and NFE2L1−/− BMDMs were subject to OGD/R followed by analysis of the indicated mitophagy regulators by WB with (J) and qRT-PCR (K). L WT and NFE2L1−/− BMDMs subject to OGD/R were analyzed for NRF1 binding to the promoters of the indicated genes by Cut&Tag-qPCR. Statistical test by two-way ANOVA (n = 3 and representative of 3 independent experiments, n.s. no significance, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001).

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