Journal: Nature Communications

Article Title: Cholangiocarcinoma PDHA1 succinylation suppresses macrophage antigen presentation via alpha-ketoglutaric acid accumulation

doi: 10.1038/s41467-025-58429-7

Figure Lengend Snippet: a Upregulated genes revealed by succinylation proteomic analysis of cholangiocarcinoma and adjacent non-tumor tissues ( n = 10). b Mass spectrometry identified the succinylation modification site K83 on PDHA1. c Quantification of PDHA1 K83Succ immunostaining performed to compare paired samples ( n = 74). d Kaplan–Meier analysis of OS curves based on PDHA1 K83Succ expression. PDHA1 K83Succ expression associated with poor prognosis in CCA patients. e Representative immunofluorescence staining of CCA and adjacent normal tissues ( n = 7). PDHA1 K83Succ (Red), DAPI (Blue). And statistical chart of PDHA1 K83Succ mean gray values. Scale bars: 50 μm. f Schematic illustrating transposon- and CRISPR-Cas9-( p53 )-based injection of vectors into mice via the tail vein to generate tumors resembling human CCA. The transposon-based vector overexpressing Kras G12D can express luciferase. Created in BioRender. Zhang, N. (2025) https://BioRender.com/y78d999 . Bioluminescence images and signal of C57BL/6 mice after plasmid injection on day 7 and day 28 ( n = 10 biologically independent samples). g Front and back representative images of livers from ( f ). Scale bars: 1 cm. h Kaplan–Meier survival curve of C57BL/6 mice treated as in ( f ). i Tumor incidence of C57BL/6 mice treated as in ( f ). j Schematic illustrating transposon- and CRISPR-Cas9-( Pten )-based injection of vectors into mice via the tail vein to generate tumors resembling human CCA. The transposon-based vector overexpressing Kras G12D can express luciferase. Created in BioRender. Zhang, N. (2025) https://BioRender.com/j69i397 . Bioluminescence images and signal of C57BL/6 mice after plasmid injection on day 7 and day 28 ( n = 8 biologically independent samples). k Front and back representative images of livers from ( j ). Scale bars: 1 cm. l Kaplan–Meier survival curve of C57BL/6 mice treated as in ( j ). m Tumor incidence of C57BL/6 mice treated as in ( j ). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t -tests ( c , e , f , j ) or log-rank test ( d , h , l ). HR hazard ratio, WT wild type. Source data are provided as a Source Data File.

Article Snippet: Anti-phospho-PDHA1-S232 mAb (81491-1-RR, 1:1000 for Western blot) and anti-phospho-PDHA1-S300 mAb (29583-1-AP, 1:1000 for Western blot) are from Proteintech.

Techniques: Mass Spectrometry, Modification, Immunostaining, Expressing, Immunofluorescence, Staining, CRISPR, Injection, Plasmid Preparation, Luciferase, Two Tailed Test

Journal: Nature Communications

Article Title: Cholangiocarcinoma PDHA1 succinylation suppresses macrophage antigen presentation via alpha-ketoglutaric acid accumulation

doi: 10.1038/s41467-025-58429-7

Figure Lengend Snippet: a tSNE (t-Distributed Stochastic Neighbor Embedding) plots of 15,830 high-quality single cells from spontaneous cholangiocarcinoma mice. b Heatmaps displaying marker genes for each cell type. c Histogram showing the proportion histogram of the eight major subtypes WT (left) and K83R (right). d tSNE plots of five different macrophage subgroups. e Dot plots showing the expression of marker genes for each cell type. f Quantitative RT-PCR analysis of mRNA levels of Cd74 , H2-Ab1 , H2-Aa , Cxcl9 , Cxcl10 changes from PDHA1 WT or PDHA1 K83R spontaneous cholangiocarcinoma mouse tumors isolated CD45 + CD11b + F4/80 + macrophages ( n = 3 biologically independent samples). g Flow cytometry analysis of CD45 + CD11b + F4/80 + MHCII + cells expression in mice with PDHA1 WT or PDHA1 K83R after hydrodynamic tail vein injection ( n = 5 biologically independent samples). h Representative mIHC staining of spontaneous cholangiocarcinoma mouse tumors. MHCII (Red), CD86 (Green), PDHA1 K83Succ (Yellow) and DAPI (Blue). Scale bars: 20 μm. i Quantification of PDHA1 K83Succ + and CD86 + MHCII + cells in PDHA1 WT and PDHA1 K83R spontaneous cholangiocarcinoma mouse tumors ( n = 5 biologically independent samples). j Representative mIHC staining of human CCA samples using antibodies against HLA-DR (Red), CD86 (Green), PDHA1 K83Succ (Yellow) and DAPI (Blue). Scale bars: 20 μm. k Pearson correlation between the levels of PDHA1 K83Succ + and CD86 + HLA-DR + in human CCA samples ( n = 20). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t -tests ( f , g , i ) or two-tailed correlation ( k ). Mat matrix, Macro macrophages, WT wild type. RNA-seq data is in the Genome Sequence Archive database and source data are provided as a Source Data File.

Article Snippet: Anti-phospho-PDHA1-S232 mAb (81491-1-RR, 1:1000 for Western blot) and anti-phospho-PDHA1-S300 mAb (29583-1-AP, 1:1000 for Western blot) are from Proteintech.

Techniques: Marker, Expressing, Quantitative RT-PCR, Isolation, Flow Cytometry, Injection, Staining, Two Tailed Test, RNA Sequencing, Sequencing

Journal: Nature Communications

Article Title: Cholangiocarcinoma PDHA1 succinylation suppresses macrophage antigen presentation via alpha-ketoglutaric acid accumulation

doi: 10.1038/s41467-025-58429-7

Figure Lengend Snippet: a Schematic diagram of THP-1 macrophages and cholangiocarcinoma cells co-culture. Created in BioRender. Zhang, N. (2025) https://BioRender.com/c42h873 . After 24 h, macrophages were used for flow analysis, and CM was used for metabolite detection ( n = 5 biological replicates). b Flow cytometry analysis of HLA-DR + MFI values after co-culturing THP-1 and the CM of QBC939 sg PDHA1 with reconstituted expression of PDHA1 WT or PDHA1 K83R after 24 h ( n = 5 biological replicates). c Heatmap representing metabolite changes in tumor WT CM compared with K83 CM was obtained after 24 h culture of tumor cells ( n = 5 biological replicates). d Schematic diagram of the TCA cycle metabolism in cancer. e Metabolite secretion of TCA cycle metabolic substrates α-KG, succinate, fumarate, malate and citrate changes in CM from QBC939 sg PDHA1 with reconstituted expression of PDHA1 WT or PDHA1 K83R. Analysis was performed after 24 h ( n = 5 biological replicates). f Flow cytometry analysis of HLA-DR + MFI values of THP-1 in co-culture with tumor CM. Analysis was performed after 24 h ( n = 5 biological replicates). g PDH enzyme activity of QBC939 sg PDHA1 with reconstituted expression of PDHA1 WT and PDHA1 K83R ( n = 5 biological replicates). Activity is represented as the rate in ΔOD/min. h Western blot analysis showed the expression of PDHA1 pS293, PDHA1 K83Succ changes in QBC939 sg PDHA1 with reconstituted expression of PDHA1 WT, PDHA1 K83R, PDHA1 S293A and PDHA1 K83R S293A. The samples derive from the same experiment but different gels for PDHA1 pS293, PDHA1 K83Succ, PDHA1 and α-Tubulin were processed in parallel. The experiment was repeated three times with similar results. i PDH enzyme activity of QBC939 sg PDHA1 with reconstituted expression of PDHA1 WT, PDHA1 K83R, PDHA1 S293A and PDHA1 K83R S293A ( n = 5 biological replicates). Activity is represented as the rate in ΔOD/min. j CM α-KG concentration of QBC939 sg PDHA1 with reconstituted expression of PDHA1 WT, PDHA1 K83R, PDHA1 S293A and PDHA1 K83R S293A ( n = 5 biological replicates). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t -tests. Mac macrophages, CM conditioned medium, MFI mean fluorescence intensity. Source data are provided as a Source Data File.

Article Snippet: Anti-phospho-PDHA1-S232 mAb (81491-1-RR, 1:1000 for Western blot) and anti-phospho-PDHA1-S300 mAb (29583-1-AP, 1:1000 for Western blot) are from Proteintech.

Techniques: Co-Culture Assay, Flow Cytometry, Expressing, Activity Assay, Western Blot, Concentration Assay, Two Tailed Test, Fluorescence

Journal: Nature Communications

Article Title: Cholangiocarcinoma PDHA1 succinylation suppresses macrophage antigen presentation via alpha-ketoglutaric acid accumulation

doi: 10.1038/s41467-025-58429-7

Figure Lengend Snippet: a Gene Ontology (GO) analysis of RNA-seq after 24 h co-culture of tumor CM with or without α-KG (8 μM) and THP-1 cells. b Volcano plot to depict the downregulated HLA-DRA , HLA-DPA1 , and HLA-DQA1 expression changes of immune response observed in the RNA-seq data. c Gene Set Enrichment Analysis (GSEA) indicating that positive regulation of MAPK cascade was significantly upregulated after α-KG accumulation. d Western blot analysis showed the expression of the MAPK pathway and HLA-DR, CD74 changes after 24 h co-culture ( n = 3 biological replicates). The samples derive from the same experiment but different gels for p-ERK, ERK, and HLA-DR, another for CD74 and p-P38, another for p-JNK, JNK, and another for P38 and α-Tubulin were processed in parallel. e Diagram of interaction between α-KG and OXGR1 protein using molecular docking. f Western blot analysis showed the expression of p-ERK, HLA-DR, and CD74 changes after 24 h co-culture ( n = 3 biological replicates). The samples derive from the same experiment but different gels for p-ERK, ERK, and HLA-DR, another for CD74, and another for OXGR1 and α-Tubulin were processed in parallel. g Schematic diagram of THP-1 with or without OXGR1 depletion and tumor CM with α-KG (0, 8 μM). Created in BioRender. Zhang, N. (2025) https://BioRender.com/a10o519 . After 24 h, macrophages were used for flow analysis. h Flow cytometry analysis of HLA-DR + MFI values from data ( g ) ( n = 4 biological replicates). i Schematic diagram of subcutaneous tumor treatment. Created in BioRender. Zhang, N. (2025) https://BioRender.com/j80q160 . And representative images of vehicle, montelukast, α-KG or combined on day 26 ( n = 5 biologically independent samples). j Tumor growth of LTP-C9 with overexpression of PDHA1 WT were injected into C57BL/6 mice ( n = 5 biologically independent samples), with vehicle, montelukast, α-KG or combined treatment, beginning on day 14. k Tumor volume from data ( i ) ( n = 5 biologically independent samples). l Tumor weight from data ( i ) ( n = 5 biologically independent samples). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t -tests ( d , f , h , k , l ) or 2way ANOVA ( j ). Each Western blot was independently repeated three times. Mac macrophages, CM conditioned medium, MFI mean fluorescence intensity. Source data are provided as a Source Data File.

Article Snippet: Anti-phospho-PDHA1-S232 mAb (81491-1-RR, 1:1000 for Western blot) and anti-phospho-PDHA1-S300 mAb (29583-1-AP, 1:1000 for Western blot) are from Proteintech.

Techniques: RNA Sequencing, Co-Culture Assay, Expressing, Western Blot, Flow Cytometry, Over Expression, Injection, Two Tailed Test, Fluorescence

Journal: Nature Communications

Article Title: Cholangiocarcinoma PDHA1 succinylation suppresses macrophage antigen presentation via alpha-ketoglutaric acid accumulation

doi: 10.1038/s41467-025-58429-7

Figure Lengend Snippet: a Immunofluorescence analysis to detect colocalization of PDHA1 and DLST in HCCC9810 and RBE cells. Scale bars: 20 μm. b Western blot analysis of input and anti-FLAG immunoprecipitates (IPs) derived from HEK293T cells transfected with FLAG-PDHA1 and HA-tagged EV, HAT1, KAT2A, CPT1A, DLST. The samples derive from the same experiment but different gels for Succ-K, another for HA, another for Flag, and another for α-Tubulin were processed in parallel. c Western blot analysis of input and anti-FLAG immunoprecipitates (IPs) derived from HEK293T cells transfected with FLAG-PDHA1-EV, FLAG-PDHA1 WT, FLAG-PDHA1 K83R, and HA-DLST. The samples derive from the same experiment but different gels for Succ-K, another for HA, another for Flag, and another for α-Tubulin were processed in parallel. d Western blot analysis of DLST changes from QBC939 cells. e Western blot analysis of PDHA1 K83Succ, PDHA1 pS293, PDHA1 changes from QBC939 cells after DLST depletion. The samples derive from the same experiment but different gels for PDHA1 K83Succ, PDHA1 pS293, and DLST, another for PDHA1, and another for α-Tubulin were processed in parallel. f PDH enzyme activity of QBC939 cells after DLST depletion ( n = 5 biological replicates). Activity is represented as the rate in ΔOD/min. g CM α-KG concentration of QBC939 cells after DLST depletion ( n = 3 biological replicates). h Flow cytometry analysis of HLA-DR + MFI values after co-culturing the CM of QBC939 cells after DLST depletion and THP-1 after 24 h ( n = 5 biological replicates). i Western blot analysis of PDHA1 pS293 and PDHA1 K83Succ changes from QBC939 sg PDHA1 with reconstituted expression of FLAG-PDHA1 WT and FLAG-PDHA1 K83R with or without DLST. The samples derive from the same experiment but different gels for PDHA1 pS293, PDHA1 K83Succ, and DLST, another for PDHA1, and another for α-Tubulin were processed in parallel. j CM α-KG concentration of QBC939 sg PDHA1 with reconstituted expression of FLAG-PDHA1 WT and Flag-PDHA1 K83R with or without DLST ( n = 5 biological replicates). k Western blot analysis of PDHA1 K83Succ, PDHA1 pS293 changes from QBC939 sgPDHA1 with reconstituted expression of FLAG-PDHA1 WT cell line used CPI-613 (0, 50, 100 μM). The samples derive from the same experiment but different gels for PDHA1 K83Succ, PDHA1 pS293, another for PDHA1 and Actin were processed in parallel. l CM α-KG concentration of QBC939 sg PDHA1 with reconstituted expression of FLAG-PDHA1 WT cell line used CPI-613 (0, 50, 100 μM) ( n = 5 biological replicates). m Flow cytometry analysis of HLA-DR + MFI values after co-culturing the CM of QBC939 sg PDHA1 with reconstituted expression of FLAG-PDHA1 WT cell line used CPI-613 (0, 50, 100 μM) ( n = 5 biological replicates). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t -tests. Each Western blot was independently repeated three times. CM conditioned medium, MFI mean fluorescence intensity. Source data are provided as a Source Data File.

Article Snippet: Anti-phospho-PDHA1-S232 mAb (81491-1-RR, 1:1000 for Western blot) and anti-phospho-PDHA1-S300 mAb (29583-1-AP, 1:1000 for Western blot) are from Proteintech.

Techniques: Immunofluorescence, Western Blot, Derivative Assay, Transfection, Activity Assay, Concentration Assay, Flow Cytometry, Expressing, Two Tailed Test, Fluorescence

Journal: Nature Communications

Article Title: Cholangiocarcinoma PDHA1 succinylation suppresses macrophage antigen presentation via alpha-ketoglutaric acid accumulation

doi: 10.1038/s41467-025-58429-7

Figure Lengend Snippet: a Schematic diagram of subcutaneous tumor treatment. Created in BioRender. Zhang, N. (2025) https://BioRender.com/g40g507 . b Tumor growth of LTP-C9 with overexpression of PDHA1 WT were injected into C57BL/6 mice ( n = 6 biologically independent samples), with vehicle, CPI-613, Gem+Cis or combined treatment, beginning on day 14, CPI-613 and vehicle were administered daily, Gem+Cis were administered on experimental days (ED) 1, 5, 9, and 13. c Representative images of tumor volumes from vehicle, CPI-613, Gem+Cis or combined treatment on day 30 ( n = 6 biologically independent samples). d Tumor volume and weight from data ( b ) ( n = 6 biologically independent samples). e Schematic diagram of spontaneous tumor treatment. Created in BioRender. Zhang, N. (2025) https://BioRender.com/v16u276 . f Bioluminescence images of C57BL/6 mice after plasmid injection on day 7, and mice on day 28 after administration of vehicle, CPI-613, Gem+Cis or combined treatment. g Bioluminescence signal from data ( e ) ( n = 10 biologically independent samples). h Front and back representative images of livers from ( e ). Scale bars: 1 cm. i Kaplan–Meier survival curve of C57BL/6 mice treated as in ( e ) ( n = 10 biologically independent samples). j Tumor incidence from data ( e ). k Flow cytometry analysis of CD45 + CD11b + F4/80 + MHCII + cells expression of vehicle, CPI-613, Gem+Cis or combined treatment ( n = 5 biologically independent samples). l Flow cytometry analysis of CD45 + CD3 + CD4 + T cells expression of vehicle, CPI-613, Gem+Cis or combined treatment ( n = 5 biologically independent samples). m Flow cytometry analysis of CD45 + CD3 + CD8 + T cells expression of vehicle, CPI-613, Gem+Cis or combined treatment ( n = 5 biologically independent samples). Data are presented as mean values ± SD, and P values were calculated using two-tailed unpaired Student’s t -tests ( d , g , k – m ), 2way ANOVA ( b ) or log-rank test ( i ). Gem gemcitabine, Cis cisplatin. Source data are provided as a Source Data File.

Article Snippet: Anti-phospho-PDHA1-S232 mAb (81491-1-RR, 1:1000 for Western blot) and anti-phospho-PDHA1-S300 mAb (29583-1-AP, 1:1000 for Western blot) are from Proteintech.

Techniques: Over Expression, Injection, Plasmid Preparation, Flow Cytometry, Expressing, Two Tailed Test

Journal: Cell reports

Article Title: ApoE4 Impairs Neuron-Astrocyte Coupling of Fatty Acid Metabolism

doi: 10.1016/j.celrep.2020.108572

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Mouse Anti-PDH-E1α Antibody , Mitoscience , Cat# MSP07; RRID: AB_478296.

Techniques: Plasmid Preparation, Recombinant, Staining, Cell Based Assay, Extraction, Software