Anti β actin

The protein expression levels of peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1α), peroxisome proliferator-activated receptor alpha (PPARα), nuclear respiratory factor 1 (NRF1), and mitochondrial transcription factor A (mtTFA) were measured by Western blot (Chen et al., 2018 (link)). The specific methods and steps are shown in Part 2 of the Supplementary Materials. The following antibodies were used: anti-PGC-1α (batch number: ab54481), anti-PPARα (batch number: ab8934), anti-NRF1 (batch number: ab175932), and anti-mtTFA (mitochondrial marker, batch number: ab131607) were provided by Abcam (Cambridge, UK); anti-β-actin (batch number: TA-09) was supplied by Beijing Zhongshan Jinqiao Biotechnology Co., Ltd. (Beijing, China). The mRNA expression levels of PGC-1α, PPARα, NRF1, and mtTFA were measured by real-time PCR (RT-PCR) (Mocker et al., 2019 (link)). The specific methods and steps are shown in Part 3 of the Supplementary Materials. Mitochondrial DNA (mtDNA) was detected using the PCR-fluorescent probe method (Liu et al., 2016 (link)). The specific methods and steps are shown in Part 4 of the Supplementary Materials.

EJ5-GFP plasmids were linearised by HindIII enzyme digestion and pCherry vectors were used to detect the
double-strand break (DSB) repair efficiency of nonhomologous end joining
(NHEJ).^{33 –35 (link)} miR-1246 mimic (sense:
5′-AAUGGAUUUUUGGAGCAGG-3′, antisense: 5′-UGCUCCAAAAAUCCAUUUU-3′); miR-NC (sense:
5′-UUCUCCGAACGUGUCACGUTT-3′, antisense: 5′-ACGUGACACGUUCGGAGAATT-3′); miR-1246
inhibitor (5′-CCUGCUCCAAAAAUCCAUU-3′); and inhibitor-NC
(5′-CAGUACUUUUGUGUAGUAGUACAA-3′) were purchased from GenePharma (Shanghai,
China).
Antibodies used in this study were as follows: anti-53BP1 (Abcam,
Cambridge, UK), anti-LIG4 (12695-1-AP; Proteintech Group Inc., Rosemont, IL, USA),
anti-CD63 (H-193, sc-15363; Santa Cruz Biotechnology, Santa Cruz, CA, USA),
anti-TSG101 (ab133586; Abcam, Cambridge, MA, USA), anti-GAPDH (TA309157; Beijing
Zhongshan Jinqiao Biotechnology Co., Ltd., Beijing, China), and anti-β-actin
(TA-09; Beijing Zhongshan Jinqiao Biotechnology Co., Ltd.).

PLA was bought from Sigma-Aldrich (St. Louis, MO, USA). And PD98059, an inhibitor of ERK1/2, was also taken from Sigma-Aldrich. PD98059 was dissolved in dimethyl sulfoxide which was purchased from Sigma-Aldrich. Fetal bovine serum (FBS) was from Gibco (Australian origin). Anti-ERK1/2 and anti-phospho-ERK1/2 were from ImmunoWay Biotechnology (Plano, TX, USA). Anti-Bax, anti-Bcl-2, and anti-caspase-3 were from Cell Signaling Technology (Danvers, MA, USA). Anti-β-actin was acquired from Zhongshanjinqiao Biotechnology (Beijing, China). Annexin V-FITC Apoptosis Detection Kit with propidium iodide (PI) double staining flow cytometry was purchased from Bestbio Biotechnology Company (Shanghai, China).

Proteins were extracted from cell lysates using RIPA lysis Buffer and the protein concentration was measured using the Enhanced BCA Protein Assay Kit (Beyotime) following to the manufacturer’s recommendations. Proteins were denatured by heat, and then separated by SDS-PAGE electrophoresis, finally transferred to PVDF membranes. The membranes were blocked using 1% bovine serum albumin solution for 1 h at room temperature, and then incubated with primary antibodies, including anti-cytochrome c (1:1000; Abcam), anti-cleaved caspase-9 (1:1000; Abcam), anti-cleaved caspase-3 (1:1000; Abcam), anti-phospho-Akt (1:1000; Abcam), anti-Akt (1:1000; Abcam), anti-phospho-Gsk-3β (1:1000; Abcam), anti-Gsk-3β (1:1000; Abcam), anti-phospho-Stat-3 (1:1000; Abcam), anti-Stat-3 (1:1000; Abcam) anti-COXIV (1:1000; Abcam) and anti-β-actin (1:1000; Zhongshan Jinqiao Biotechnology, Beijing, China) at 4°C overnight, followed by incubation with HRP-conjugated secondary antibody (1:5000; Zhongshan Jinqiao Biotechnology, Beijing, China) at room temperature for 30 min. Protein bands were detected using a BeyoECL Plus Kit (Beyotime) following to the manufacturer’s protocols. Relative densitometry was analyzed using Image J2x analysis software (NIH, U.S.A.).