Fig 1: TAZ stimulates exercise-induced mitochondrial biogenesis and increases respiratory metabolism and activity.a Wild-type (WT) and muscle-specific TAZ knockout (mKO) mice were subjected to endurance exercise training. The gastrocnemius muscle obtained from these mice was isolated and embedded in paraffin. The sectioned tissues were immunostained with an anti-COX4 antibody to visualise the mitochondria. Scale bar = 50 μm. b Genomic DNA was isolated from the gastrocnemius muscle of mice described in (a). Relative mitochondrial DNA copy number was determined via quantitative PCR using primers for mitochondrial-encoded Cox2 and nuclear-encoded β-globin. The Ct values of mitochondrial Cox2 were normalized to those of nuclear β-globin; n = 6 for each genotype (WT untrained vs. mKO untrained; *p = 0.0188, WT untrained vs. WT trained; **p = 0.0069, WT trained vs. mKO trained; ***p = 0.0002). c Protein from the gastrocnemius muscle of mice described in (a) was assessed via immunoblotting to observe the indicated proteins. GAPDH was used as the loading control. d RNA was isolated from the gastrocnemius muscle of mice described in (a). Mitochondrial-encoded marker gene expression was analysed via quantitative reverse transcription (qRT)-PCR, n = 6 for each genotype (mt-Atp6; **p = 0.0035 for WT untrained vs. mKO untrained, **p = 0.0071 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained, mt-Co2; *p = 0.0174 for WT untrained vs. mKO untrained, ****p < 0.0001 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained, mt-Cytb; *p = 0.0214 for WT untrained vs. mKO untrained, *p = 0.0124 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained, mt-Nd5; *p = 0.0186 for WT untrained vs. mKO untrained, **p = 0.0059 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained). e Nuclear-encoded mitochondrial gene expression in the gastrocnemius muscle of mice in (a) was analysed via qRT-PCR, n = 6 for each genotype (Atp5a1; *p = 0.0137 for WT untrained vs. WT trained, **p = 0.0048 for eKO untrained vs. eKO trained, Idh3g; **p = 0.0089 for WT untrained vs. WT trained, Ndufa10; *p = 0.0205 for WT untrained vs. WT trained, Uqcrc1; ****p < 0.0001 for WT untrained vs. WT trained, **p = 0.0011 for mKO untrained vs. mKO trained). f The respiratory metabolism of mice described in (a) was analysed using the Oxylet system. Data are shown as the average values observed under light and dark conditions; n = 6 for each condition (VO2-light; **p = 0.0033 for WT untrained vs. mKO untrained, **p = 0.0098 for WT trained vs. mKO trained, VO2-dark; *p = 0.0146 for WT untrained vs. mKO untrained, ***p < 0.0007 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained, VCO2-dark; ****p < 0.0001 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained, Energy expenditure-light; *p = 0.0234 for mKO untrained vs. mKO trained, Energy expenditure-dark; *p = 0.021 for WT untrained vs. mKO untrained, ***p = 0.0001 for WT untrained vs. WT trained, ****p < 0.0001 for WT trained vs. mKO trained). Ten to twelve-week-old mice were used for all panels. Data are presented as mean ± SEM for (b, d–f). Statistical significance was analysed via two-way ANOVA with Sidak’s multiple comparison test. Representative data was shown and experiments were performed at least twice with similar results for (a, c). Source data are provided as a Source data file.
Fig 2: AIF localization in the trauma hemorrhage induced CD murine model. (A) AIF expression as demonstrated by western blot analysis. The total AIF expression is similar in the Sham and THS models (0.20 vs. 0.17, respectively). (B) Cellular compartmentalization of AIF as demonstrated by WB analysis. A higher proportion of AIF is found outside the mitochondria in the THS models. The mean ratio of mitochondrial AIF (mAIF) to supernatant AIF (sAIF) is 1.5 in sham compared to 0.4 in THS group, p = 0.16. When comparing total AIF which includes the nucleus, a lower proportion is still found in the mitochondria. Ratio of mitochondrial AIF (mAIF) to total AIF (tAIF) in sham group is 0.6 compared to 0.3 in THS animals, *p = 0.04. This is suggestive of its translocation of the AIF out of the mitochondrion, potentially to the nucleus, triggering initiation of cells death. IHC sections from TH animals, DAPI (blue), AIF (green), MTCO2 (red), and combined images. In the combined images, localization of AIF in the nucleus can be seen in the THS (D) which in absent in the sham animals (C).
Fig 3: Placing COA4 in the mitochondrial copper delivery pathway to cytochrome c oxidase through the identification of its physical interactor.a A schematic representation of the mitochondrial copper delivery pathway to cytochrome c oxidase (CcO). Copper delivery to COX1 and COX2 subunits of CcO requires metallochaperones—COX17, SCO1, and COX11—that bind and transfer copper to their target proteins, and accessory factors such as COX16, COX19, SCO2, and COA6 that enable this transfer. Intermembrane space (IMS) localized CcO assembly factors depicted in yellow circles are hypothesized to play a role in copper delivery. b A heatmap of ipTM scores of the indicated human and c yeast protein pairs that are either a part of the mitochondrial copper delivery pathway or are implicated in the copper delivery process. The color bar represents the mean ipTM score. d Predicted structure of COA4-COX11 interaction. e A volcano plot of the relative abundance of proteins immunoprecipitated from V5-tagged Coa4 yeast mitochondria compared to untagged mitochondria versus statistical significance (n = 3 biological replicates). Statistical analysis was performed using a two-sided Student’s t test. f Co-immunoprecipitation of COX11-FLAG (n = 3) and g COA4-V5 from digitonin solubilized mitochondria of 293T cells transfected with COA4-V5 and COX11-FLAG followed by western blotting (n = 3). h SDS-PAGE/western blot analysis of COA4 from WT and COA4-KO MCH58 cell lines. ATP5A is used as a loading control (n = 2). i SDS-PAGE/western blot analysis of COX11 in solubilized mitochondria isolated from WT and COA4-KO-6 MCH58 cells (n = 3). j Mitochondrial Cu levels measured in WT and COA4-KO-6 mitochondria using inductively coupled plasma–mass spectrometry (n = 3). Data are presented as mean ± SD. Statistical analyses were performed using an unpaired two-tailed Student’s t test. k SDS-PAGE/western blot analysis of COX1 from WT and COA4-KO-6 cells treated with different concentrations of the copper chelator BCS (n = 3). l BN-PAGE/western blot analysis of WT and COA4-KO-6 mitochondria isolated from cells grown with and without BCS (n = 2). m Oxygen consumption rate measurements of WT and COA4-KO-6 MCH58 cells following treatment with ATP synthase inhibitor (Oligomycin), uncoupler (CCCP), and electron transport inhibitor (Antimycin A) (n = 3). Data are presented as mean ± SEM. Statistical analyses were performed using ordinary one-way ANOVA. All sample sizes in this figure represent biological replicates from independent experiments. Statistical significance levels: **p < 0.01; ****p < 0.0001. Western blots are representative of the indicated number of independent biological replicates. Source data are provided as a Source Data file. OMM outer mitochondrial membrane, IMM inner mitochondrial membrane.
Fig 4: Using AlphaFold-Multimer as a classification model to predict mitochondrial protein-protein interactions.a A schematic showing the application of AlphaFold-Multimer as a classifier to predict protein-protein interactions based on its output: the interface predicted template modeling (ipTM) score and follow-up experimental validation. Parts of this image were created in BioRender. Swaminathan, A. (2026) https://BioRender.com/lkvpwsc and Swaminathan, A. (2026) https://BioRender.com/l0d23t7. b A plot of the number of protein-protein interactions that fall into different ipTM bins for the recent PDB dataset. Solid line represents the mean ipTM score, and dotted line represents the best ipTM score from five different predicted models of interactions. The interacting and non-interacting groups have sample sizes of 1338 and 15,005 protein pairs, respectively. c Precision-recall curve of b at a 1:100 ratio of true:false pairs. The shaded part of the curve indicates a confidence interval of 95%. d Interaction network of predicted mitochondrial protein-protein interactions. Proteins are represented as circles, and the interactions are represented as gray lines. The thickness of lines is proportional to the mean ipTM scores. Blue dots represent proteins with a MitoCarta pathway annotation, and black dots represent uncharacterized proteins. e A pie chart of the predicted mitochondrial interactions mapped to known databases. Predicted structures of the f mitochondrial processing peptidase complex (PMPCB-PMPCA), g glutamyl-t-RNA(Gln) amidotransferase complex (GATA-GATB-GATC), h interaction between Fe-S cluster biogenesis factors (BOLA3-GLRX5), and i cytochrome c oxidase assembly intermediate (COX2-COX20). j A heat map depicting the predicted interactions for uncharacterized mitochondrial proteins across different MitoCarta Pathways. Predicted structures of k TCAIM-OGDH, l TCAIM-HSPA9, m C16ORF91-UQCC1, and n C16ORF91-MT-CYB interactions. Source data are provided as a Source Data file. Co-IP co-immunoprecipitation.
Supplier Page from Abcam for Anti-MTCO2 antibody