Journal: eLife

Article Title: Tissue-specific mitochondrial HIGD1C promotes oxygen sensitivity in carotid body chemoreceptors

doi: 10.7554/eLife.78915

Figure Lengend Snippet: Reagents and resources.

Article Snippet: COX4I1 -Myc-FLAG in pCMV6-Entry , OriGene , RC209374.

Techniques: Sequencing, In Situ Hybridization, Plasmid Preparation

Journal: Metabolites

Article Title: Effect of Expression of Nuclear-Encoded Cytochrome C Oxidase Subunit 4 Isoforms on Metabolic Profiles of Glioma Cells

doi: 10.3390/metabo12080748

Figure Lengend Snippet: Metabolite profiling of U251 glioma cells overexpressing the COX4-1 or COX4-2 isoform. ( A ) Two-dimensional PCA score plots of untargeted metabolomics data from COX4-1-overexpressing glioma cells (blue) and COX4-2-overexpressing glioma cells (red). ( B ) Volcano plot of metabolomic data. The x-axis represents the mean fold change (log 2 ratio) in the relative intensity of each metabolite between the two samples (COX4-2-overexpressing relative to the COX4-1-overexpressing cells). The y-axis represents the statistical significance (-log 10 -transformed p -values) of each metabolite. Metabolites are color coded by metabolic pathway.

Article Snippet: U251 COX4-2-KO cells were electroporated with CMV6 plasmids containing FLAG-epitope-tagged COX4-2 or COX4-1 (Catalog # RC209204 and RC209374, OriGene Technologies, Rockville, MD, USA).

Techniques: Transformation Assay

Journal: Nature Communications

Article Title: Lactate mitochondrial oxidation drives stemness potential in metastatic breast cancer

doi: 10.1038/s41467-025-67091-y

Figure Lengend Snippet: A , B Schematic of 2- 2 H-lactate metabolism ( A ) and 2 H-labeled M + 1 metabolites ( B , n = 3 biologically independent samples). ** P = 0.0018, **** P < 0.0001; two-tailed t test. C , D CD147/MCT1/LDHB protein levels in mitochondria ( C ) and sub-compartments ( D ) from AR cells. K proteinase K, T triton X-100, O/I outer membrane and intermembrane space, I/M inner membrane and matrix. COX IV, Na + -K + ATPase, α-Tubulin, TOMM20 mark of inner membrane, plasma membrane, cytoplasm, outer membrane. Data are representative of 3 independent experiments. E Transmission electron microscopy shows CD147/MCT1/LDHB co-localization (orange arrows) in the mitochondrial inner membrane of AR cells. Scale bars: 1 μm (left), 500 nm (right). Red asterisks, mitochondria. F Proximity ligation assay (PLA) between CD147 and LDHB in lactate-treated MCF7 cells. Scale bars, 5 μm. n = 6 typical fields from 3 biologically independent samples. DAPI (blue), PLA signal (red). **** P < 0.0001; two-tailed t test. G Pull-down assay between GST-CD147 N-terminal and His-LDHB. Representative of 3 independent experiments. H CD147 (red)/LDHB (blue)/MCT1 (purple) co-localization (orange arrows) in COX IV (green) labeled mitochondria from PDX1 lung metastatic tumors. Scale bars: 20 μm. I CD147 (green)/LDHB (yellow)/MCT1 (orange) expressions in CK18 (red) labeled cells from breast cancer patients with or without metastasis. Scale bars: 40 μm. J Correlation between CD147 and MCT1/LDHB in 11 metastatic patients with breast cancer. R 2 , Pearson correlation coefficient. Spearman correlation analysis. K Kaplan–Meier survival (up) and recurrence (down) analysis for 131 patients stratified by CD147/MCT1/LDHB co-expression (high n = 9 patients, low n = 31 patients). L Representative images of HE (left: scale bars, 50 μm) and SOX2 IHC staining (middle: scale bars, 50 μm ; right : enlarged image, scale bars, 10 μm) in CTCs from No. 9 patient in Fig. . M Co-localization (orange arrows) of CD147 (red)/LDHB (blue)/MCT1 (purple) in COX IV (green) labeled mitochondria of CTCs from No. 9 patient in Fig. . Scale bars: 20 μm. N SOX2 IHC intensity in CTCs from 10 patients stratified by mitochondrial CD147/MCT1/LDHB complex presence (without, n = 4 patients; with, n = 6 patients). *** P = 0.0004, two-tailed t test. Data are mean ± SEM. Source data are provided as a Source Data file.

Article Snippet: For cellular co-localization analysis among CD147, MCT1, LDHB, and COX IV, cells were initially stained with primary antibodies against CD147 (1:200), MCT1 (1:200; 20139-1-AP, Proteintech), and LDHB (1:100; ab195964, Abcam) overnight at 4 °C, followed by incubation with Alexa Fluor 555-, 647- or 405-conjugated antibodies, respectively, at room temperature for 1 h. This was followed by incubation with an Alexa Fluor 488-conjugated COX IV antibody (1:200; CL488-60251, Proteintech) at room temperature for 3–4 h. Finally, nuclei were counterstained, and images were captured following the same procedures as described above.

Techniques: Labeling, Two Tailed Test, Membrane, Clinical Proteomics, Transmission Assay, Electron Microscopy, Proximity Ligation Assay, Pull Down Assay, Expressing, Immunohistochemistry

Journal: Nature Communications

Article Title: Lactate mitochondrial oxidation drives stemness potential in metastatic breast cancer

doi: 10.1038/s41467-025-67091-y

Figure Lengend Snippet: A Immunoactivity of EGFP-fused mito-CD147 scFv to CD147 in MCF7/T47D cell subfractions: C (plasma membrane and cytoplasm), M (mitochondria), COX IV, α-Tubulin mark mitochondrial inner membrane, cytoplasm. NC: pGV469 vector; scFv: pGV469-mito-CD147 scFv. Representative of 3 independent experiments. B Co-IP assays showing CD147 interaction with MCT1/LDHB in mitochondria from lactate-treated MCF7 WT/KO/mito147 cells as indicated. Groups: 1. WT; 2. WT+mito-CD147 scFv; 3. KO; 4. KO+mito-CD147 scFv; 5. mito147; 6. mito147+mito-CD147 scFv. Representative of 3 independent experiments. C–F Experimental design ( C ), lung nodules ( D , n = 10 mice/group), ALDH + and CD44 high CD24 −/low ratios ( E , n = 5 mice/group), and SOX2 levels ( F , n = 4 mice/group, α-Tubulin, loading control) in tumors from MMTV-PyMT mice as indicated. *** P = 0.0001 ( E left), *** P = 0.0004, 0.0003 ( E right, NS vs Lac, Lac vs Lac + scFv), **** P < 0.0001, one-way ANOVA with Bonferroni correction. C created in BioRender. Song, C. (2025) https://BioRender.com/37z58my . G , H CTC counts ( G , left, n = 8 mice/group), SOX2 intensity ( G , right; NS, n = 148 cells from 7 mice; Lac, n = 342 cells from 7 mice; Lac+scFv, n = 149 cells from 7 mice), and representative images of CTCs ( H ; scale bars, 40 μm), from MMTV-PyMT mice as indicated. H DAPI (blue), CK18 (green), SOX2 (purple). **** P < 0.0001, one-way ANOVA with Bonferroni correction. I , J Lung nodules ( I , n = 8 mice/group), ALDH + and CD44 high CD24 −/low ratios ( J , n = 5 mice/group) in tumors from PDX1-bearing mice as indicated. ns, P = 0.9818, 0.869 ( J left, right), *** P = 0.0001, **** P < 0.0001, one-way ANOVA with Bonferroni correction. K–M CTC counts ( K , left, n = 8 mice/group), SOX2 intensity ( K , right; NS, n = 80 cells from 8 mice; scFv, n = 80 cells from 8 mice; Lac, n = 120 cells from 6 mice; Lac+scFv, n = 80 cells from 8 mice), representative CTC images ( L ), and mito-CD147 scFv binding to mitochondrial CD147 ( M ) in CTCs from PDX1-bearing mice as indicated. Scale bars: 10 μm. L DAPI (blue), CK18 (green), SOX2 (purple). M DAPI (blue), CD147 (red), scFv (green), COX IV (purple). CTC circulating tumor cell, scFv single-chain variable fragment. Data are presented as mean ± SEM. Source data are provided as a Source Data file.

Article Snippet: For cellular co-localization analysis among CD147, MCT1, LDHB, and COX IV, cells were initially stained with primary antibodies against CD147 (1:200), MCT1 (1:200; 20139-1-AP, Proteintech), and LDHB (1:100; ab195964, Abcam) overnight at 4 °C, followed by incubation with Alexa Fluor 555-, 647- or 405-conjugated antibodies, respectively, at room temperature for 1 h. This was followed by incubation with an Alexa Fluor 488-conjugated COX IV antibody (1:200; CL488-60251, Proteintech) at room temperature for 3–4 h. Finally, nuclei were counterstained, and images were captured following the same procedures as described above.

Techniques: Clinical Proteomics, Membrane, Plasmid Preparation, Co-Immunoprecipitation Assay, Control, Binding Assay