Anti sdha

Blue native gel electrophoresis (PAGE) was performed by isolating mitochondrial proteins from MT and WT zebrafish, as detailed previously (50 (link)). Samples containing 15 μg of proteins were separated on 3–11% Bis-Tris Native PAGE gel. The primary antibodies applied for this experiment were anti-Ndufa9 (Proteintech, 20312-1-AP), anti-Sdha (Proteintech, 14865-1-AP), anti-Uqcrc2 (Abcam, ab203832), anti-Cox5a (Proteintech, 11448-1-AP), anti-Atp5c (Proteintech 60284-1-Ig) and anti-Vdac1/2(Proteintech, 10866-1-AP). Peroxidase AffiniPure goat anti-mouse IgG and goat anti-rabbit IgG (Jackson) were used as secondary antibodies and protein signals were detected using the ECL system (CWBIO).

Western blotting analysis was performed as detailed elsewhere (42 (link),44 (link),49 (link)). Fish were sacrificed after anesthesia and homogenized in RIPA reagent (Invitrogen) using a homogenizer. Twenty micrograms of total cellular proteins were electrophoresed through 10% bis-Tris SDS-polyacrylamide gels and then transferred to a polyvinyl difluoride (PVDF) membrane. The antibodies used include anti-Gtpbp3 (Sigma, HPA042158), anti-Nd5 (Abcam, ab92624), anti-Nd6 (Abcam, ab81212), anti-Cytb (ABclonal, A9762), anti-Co2 (Proteintech, 55070-1-AP), anti-Sdha (Proteintech, 14865-1-AP), anti-Atp5c (Proteintech 60284-1-Ig), anti-Tfam (Proteintech, 19998-1-AP), and anti-Tufm (Proteintech, 26730-1-AP), anti-Yars2 (Abcam, ab228957), anti-Kars (Proteintech, 14951-1-AP), anti-Afg3l2 (Proteintech, 14631-1-AP), anti-Clpp (Proteintech, 15698-1-AP) and anti-Gapdh (Sigma, SAB2701826). Peroxidase AffiniPure goat anti-mouse IgG and goat anti-rabbit IgG (Jackson) were used as secondary antibodies and protein signals were detected using the ECL system (CWBIO). Quantification of density in each band was performed as detailed previously (42 (link)).

Protein extraction and WB analysis were carried out following the manufacturer's protocol. The primary antibodies used in this study included: anti-H3K27me3 (CST, Cat# 9733); anti-RNA polymerase II (Sigma-Aldrich, Cat# 05-623); anti-ATF3 (CST, Cat# 18665); anti-SDHA (Protein-tech, Cat# 14865-1-AP); anti-β-actin (CST, Cat# 3700); anti-β-tubulin (CST, Cat# 2146); anti-p21 Waf1/Cip1 (CST, Cat# 2947); anti-CDK4 (Affinity Biosciences, Cat# DF6102); anti-CDK6 (Abcam, Cat# ab124821); anti-cyclin D1 (Affinity Biosciences, Cat# AF0931); anti-Retinoblastoma (Rb) (Affinity Biosciences, Cat# DF6840); anti-Phospho-Retinoblastoma (p-Rb) (Affinity Biosciences, Cat# AF3103); anti-cyclin E1 (Affinity Biosciences, Cat# AF0144); anti-cyclin A (Affinity Biosciences, Cat# AF0142); anti-HK2 (CST, Cat# 2867). After probing with primary antibodies, corresponding horseradish peroxidase (HRP)-conjugated secondary antibodies were used to visualize the protein bands.