Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A Relative MRPL13 mRNA expression in the GEPIA database. Box plots show MRPL13 mRNA expression in OC ( n = 426) and the corresponding normal tissues ( n = 88). Axis unit is log2(TPM + 1). B Relative MRPL13 mRNA expression in OC and normal tissues based on data from the GEO database ( GSE26712 , GSE54388 , GSE18520 , GSE27651 and GSE36668 ). C Relative MRPL13 protein expression in OC ( n = 100) and normal tissues ( n = 25) based on data from the UALCAN database. D Representative images of MRPL13 IHC staining in ovarian malignant tumors ( n = 114), ovarian borderline tumors ( n = 33), ovarian benign tumors ( n = 31) and ovarian normal tissues ( n = 13). Scale bar, 100 μm (×200, upper) and 50 μm (×400, lower). E Scatter plot of MRPL13 IHC staining scores in different ovarian tissues. F Kaplan–Meier analysis of the OS of patients with OC in the Shengjing hospital cohort based on MRPL13 expression evaluated in. G , H Kaplan–Meier analysis of the OS and PFS of patients with OC in Kaplan–Meier plotter database based on MRPL13 expression. I , J Forest plots of univariate and multivariate Cox regression analyses for various clinicopathological parameters in OC patients. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: Expressing, Immunohistochemistry

Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A CCK-8 assays were utilized to evaluate the cell viability in OVCAR-3 and ES-2 cells with MRPL13 knockdown. *: siControl vs siMRPL13. B EdU assays were conducted to assess cell proliferation in OVCAR-3 and ES-2 cells with MRPL13 knockdown. DAPI (blue) staining was performed to indicate total cells, while EdU (red) incorporation indicated cells with active DNA replication. C Transwell assays were conducted to assess cell invasion ability in OVCAR-3 and ES-2 cells with MRPL13 knockdown. D Wound healing assays were used to evaluate cell migration ability in OVCAR-3 and ES-2 cells with MRPL13 knockdown. E Cell apoptosis of OVCAR-3 and ES-2 cells in the MRPL13 knockdown groups was validated by flow cytometry. F Protein expression level of PCNA, Bcl-2, Bax, Caspase-3 and Cleaved Caspase-3 were monitored by western blot for lysates from OVCAR-3 and ES-2 cells with MRPL13 knockdown. β-Actin was used as an internal control. All assays were performed in three independent experiments. G Subcutaneous xenograft tumor model of OC was established in BALB/c nude mice with MRPL13 stable overexpression ES-2 cells. H The weights of subcutaneous xenograft tumors were measured after euthanized. I Tumor volumes were measured at three-day intervals following the subcutaneous injection to generate tumor growth curve. J Representative images of Hematoxylin-eosin (H&E) staining and IHC staining on the paraffin sections of xenograft tumors to visualize the expression of Ki-67 and MRPL13. Scale bar, 100 μm (×200, upper) and 50 μm (×400, lower). Data are presented as mean ± SD. **, P < 0.01; ***, P < 0.001.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: CCK-8 Assay, Knockdown, Staining, Migration, Flow Cytometry, Expressing, Western Blot, Control, Over Expression, Injection, Immunohistochemistry

Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A , B Bubble plots of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis results for the co-expressed genes of MRPL13 based on David database. C Results of GSEA comparing the enrichment of the oxidative phosphorylation pathway between patients with high MRPL13 expression and low MRPL13 expression in TCGA-OV cohort. D OCR was measured in OVCAR-3 and ES-2 cells with MRPL13 knockdown using Seahorse XF96 Extracellular Flux Analyzer. Basal respiration, maximal respiration and ATP production related OCR were quantitatively calculated. E Relative ATP level was recorded in OVCAR-3 and ES-2 cells with MRPL13 knockdown by ATP assay kit. F The mean fluorescence intensity (MFI) of DCF-DA was quantitatively calculated by flow cytometry to assess the ROS level in OVCAR-3 and ES-2 cells with MRPL13 knockdown. G Mitochondrial membrane potential in OVCAR-3 and ES-2 cells with MRPL13 knockdown was determined by quantitative analysis of the MFI of JC-1 aggregates (red)/monomers (green). H The degree of mPTP opening in OVCAR-3 and ES-2 cells with MRPL13 knockdown was detected by quantitative analysis of the MFI of Calcein AM fluorescent probe. I Fluorescence staining (Mito-Tracker Red CMXRos, red; Hoechst, blue) showing the mitochondrial morphology in OVCAR-3 and ES-2 cells with MRPL13 knockdown. The mitochondrial length was quantitatively analyzed by ImageJ Mitochondria Analyzer. J Representative images of TEM showing the mitochondrial morphology in OVCAR-3 and ES-2 cells with MRPL13 knockdown. All assays were performed in three independent experiments. Data are presented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: Phospho-proteomics, Expressing, Knockdown, ATP Assay, Fluorescence, Flow Cytometry, Membrane, Staining

Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A Proteins that interacted with MRPL13 were identified by combining Co-IP and mass spectrometry. The images of coomassie brilliant blue staining were displayed, and the locations of target proteins were marked with black arrows. B Venn diagram shows the number of proteins interacting with MRPL13 or IgG in mass spectrometry result. C Secondary mass spectrum of SLC25A6 peptide interacting with MRPL13 from the results of mass spectrometry. D Binding between endogenous MRPL13 and SLC25A6. Lysates of OVCAR-3 cells were immunoprecipitated in the presence of control IgG and with anti-MRPL13 or anti-SLC25A6 antibodies. Immunoblotting was performed to detect the MRPL13 and SLC25A6 precipitates using anti-SLC25A6 and anti-MRPL13 antibodies, respectively. E Binding between exogenous MRPL13 and SLC25A6. HEK293T cells were transiently transfected with plasmids encoding either Flag-MRPL13 or HA-SLC25A6 alone or in combination. Cell lysates were immunoprecipitated in the presence of anti-Flag or anti-HA antibodies. Immunoblotting was performed using anti-HA and anti-Flag antibodies, respectively. F Molecular docking of the 3D structure revealed the tight interaction between MRPL13 (purple) and SLC25A6 (blue). G Visualization of MRPL13 and SLC25A6 endogenous interaction. OVCAR-3 and HEK293T cells were analyzed by IF staining for detecting the localization of MRPL13 (red) and SLC25A6 (green). DAPI was used as a nuclear stain (blue). H Visualization of MRPL13 and SLC25A6 exogenous interaction. OVCAR-3 and HEK293T cells were co-transfected with plasmids encoding Flag-MRPL13 and Myc-SLC25A6 and analyzed by IF staining for detecting the localization of Flag (red) and Myc (green). DAPI was used as a nuclear stain (blue). I Schematic representation of the domain of full-length (FL) MRPL13 or SLC25A6 and different truncation mutants. J Lysates from HEK293T cells were transiently transfected with HA-SLC25A6 and FLAG-tagged FL or truncated MRPL13 constructs and immunoprecipitated with anti-HA antibody, followed by immunoblotting with anti-Flag and anti-HA antibodies. K Lysates from HEK293T cells were transiently transfected with Flag-MRPL13 and Myc-tagged FL or truncated SLC25A6 constructs and immunoprecipitated with anti-Flag antibody, followed by immunoblotting with anti-Myc and anti-Flag antibodies. All assays were performed in three independent experiments.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: Co-Immunoprecipitation Assay, Mass Spectrometry, Staining, Binding Assay, Immunoprecipitation, Control, Western Blot, Transfection, Construct

Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A , B The protein and mRNA expression levels of SLC25A6 were measured by western blot and qRT-PCR in OVCAR-3 and ES-2 cells with MRPL13 knockdown or overexpression, respectively. β-Actin was used as an internal control. C Stability analysis of SLC25A6 protein half-life was validated by western blot in MRPL13-knockdown OVCAR-3 cells after treatment with 20 μg/mL CHX for the indicated times. D Stability analysis of SLC25A6 protein half-life was validated by western blot in OVCAR-3 cells after treatment with 20 μg/mL CHX and either 10 μM MG132 or 400 nM BafA1 for the indicated times. E , F SLC25A6 protein half-life plots were derived by quantifying the relative levels of SLC25A6 to β-actin protein, based on band intensity. G , H Ubiquitination assay of SLC25A6 in OVCAR-3 and ES-2 cells with MRPL13 overexpression treated with 10 μM MG132 for 6 h. Cell lysates were immunoprecipitated in the presence of anti-SLC25A6 antibodies. Immunoblotting was performed using anti-Ubiquitin and anti-SLC25A6 antibodies. I Ubiquitination assay of SLC25A6 in HEK293T cells co-transfected with HA-Ub, Myc-SLC25A6 or Flag-MRPL13 and treated with 10 μM MG-132 for 6 h. Cell lysates were immunoprecipitated in the presence of anti-Myc antibodies. Immunoblotting was performed using anti-HA and anti-Myc antibodies. J Ubiquitination assay of SLC25A6 in HEK293T cells co-transfected with Myc-SLC25A6, Flag-MRPL13 or the HA-tagged K6-, K11-, K27-, K29-, K33-, K48-, or K63-only ubiquitin mutant (for example, K6 indicates that all lysines with the exception of K6 were mutated to arginine) and treated with 10 μM MG-132 for 6 h. Cell lysates were immunoprecipitated in the presence of anti-Myc antibodies. Immunoblotting was performed using anti-HA and anti-Myc antibodies. All assays were performed in three independent experiments. Data are presented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Knockdown, Over Expression, Control, Derivative Assay, Ubiquitin Proteomics, Immunoprecipitation, Transfection, Mutagenesis

Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A CCK-8 assays were utilized to evaluate the cell viability in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. *: Vector vs SLC25A6; #: MRPL13 vs MRPL13 + SLC25A6. B EdU assays were conducted to assess cell proliferation in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. DAPI (blue) staining was performed to indicate total cells, while EdU (red) incorporation indicated cells with active DNA replication. C Transwell assays were conducted to assess cell invasion ability in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. D Wound healing assays were used to evaluate cell migration ability in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. E Cell apoptosis of OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression was validated by flow cytometry. F Protein expression level of PCNA, Bcl-2, Bax, Caspase-3 and Cleaved Caspase-3 were monitored by western blot for lysates from OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 knockdown. β-Actin was used as an internal control. All assays were performed in three independent experiments. Data are presented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: CCK-8 Assay, Over Expression, Plasmid Preparation, Staining, Migration, Flow Cytometry, Expressing, Western Blot, Knockdown, Control

Journal: Cell Death & Disease

Article Title: MRPL13 enhances mitochondrial function and promotes tumor progression in ovarian cancer by inhibiting mPTP opening via SLC25A6

doi: 10.1038/s41419-025-07953-x

Figure Lengend Snippet: A The degree of mPTP opening in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression was detected by quantitative analysis of the MFI of Calcein AM fluorescent probe. B Distribution of Cyt c was detected in cytoplasm (Cyto) and mitochondrial (Mito) lysate by western blot. Lysates were collected from OVCAR-3 cells with MRPL13 and/or SLC25A6 knockdown. β-Actin was used as an internal control in cytoplasm. COX IV was used as an internal control in mitochondrial. C Mitochondrial membrane potential in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression was determined by quantitative analysis of the MFI of JC-1 aggregates (red)/monomers (green). D The mean fluorescence intensity (MFI) of DCF-DA was quantitatively calculated by flow cytometry to assess the ROS level in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. E Fluorescence staining (Mito-Tracker Red CMXRos, red; Hoechst, blue) showing the mitochondrial morphology in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. The mitochondrial length was quantitatively analyzed by ImageJ Mitochondria Analyzer. F Representative images of TEM showing the mitochondrial morphology in OVCAR-3 and ES-2 cells with MRPL13 and/or SLC25A6 overexpression. G Schematic depicting the molecular mechanism by which MRPL13 improves mitochondrial function and facilitates OC progression by inhibiting mPTP opening through SLC25A6. All assays were performed in three independent experiments. Data are presented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: The primary antibodies used were MRPL13 (Proteintech, 16241-1-AP, 1:100) and Ki-67 (CST, 9449, 1:2000).

Techniques: Over Expression, Western Blot, Knockdown, Control, Membrane, Fluorescence, Flow Cytometry, Staining