Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: Mice were treated with AOM/DSS, and at days 0 (Healthy), 15 (Colitis) and 60 (Cancer), CD4 + and CD8 + T cells were sorted from mLN and their proteomes analysed by quantitative mass spectrometry. (A) Total mLN cell number, (B) Total T cell number (left), CD4 + (centre) and CD8 + T cells (right) in the mLN of healthy, colitis and CAC mice. Each dot represents a single mouse (n=4-5). (C) Total protein content in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (D-E) Volcano plots showing the difference in the expression of proteins between (D) Colitis and Healthy CD4 + T cells and (E) Cancer and Healthy CD4 + T cells. The horizontal lines indicate a p -value of 0.05. The vertical lines indicate a fold change of ± 1.5. The percentage and number of proteins that are downregulated (blue), upregulated (red), or unchanged (grey) are indicated. (F) Venn diagrams showing the overlaps of proteins upregulated or downregulated in CD4 + T cells from mLN over Colitis and Cancer relative to Healthy. (G) Bubble plot showing the enrichment analysis of GO biological processes of the differentially upregulated (left) or downregulated (right) proteins in Cancer versus Healthy in CD4 + T cells. Bubble size indicates the number of genes involved in each process, and bubble colour reflects the fold-change in Cancer over healthy conditions. (H) Total protein content in CD8 + T cells along CAC progression. Each dot represents a pool of three mice (n=3 pools). (I-J) Volcano plots showing the difference in the expression of proteins between (I) Colitis and Healthy CD8 + T cells and (J) Cancer and Healthy CD8 + T cells. The horizontal lines indicate a p -value of 0.05. The vertical lines indicate a fold change of ± 1.5. The percentage and number of proteins that are downregulated (blue), upregulated (red), or unchanged (grey) are indicated. (K) Venn diagrams showing the overlap of proteins upregulated or downregulated in CD8 + T cells Colitis and Cancer relative to Healthy. (L) Bubble plot showing the enrichment analysis of GO biological processes of the differentially upregulated (left) or downregulated (right) proteins in Cancer versus Healthy in CD8 + T cells. Bubble size indicates the number of genes involved in each process, and bubble colour reflects the fold-change in Cancer versus Healthy conditions. All the data are shown as mean ± SEM. In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Mass Spectrometry, Expressing

Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: Mice were treated with AOM/DSS, and at days 0 (Healthy), 15 (Colitis) and 60 (Cancer), CD4 + T cells were sorted from mLN and their proteomes analysed by quantitative mass spectrometry. (A) Total ribosome mass in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (B-D) Heat maps of proteins involved in (B) ribosomal proteins (GO:0005840), (C) regulation of protein synthesis machinery and (D) amino acid transporters (GO:0006865) identified in CD4 + T cells proteomes during CAC progression. BR, biological replicate. Relative protein abundance is graded from low (blue) to high (red) to allow comparisons between different CAC stages. (E) Protein copy numbers per cell (Molecules (n°)) of the amino acid transporters SLC3A2, SLC38A1 and SL7A5 in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (F) Schematic representation of translation initiation complex. Image created with BioRender. (G-H) Stoichiometric models for (G) EIF4 and (H) EIF2 complexes. (G) Protein copy numbers for PABPC1, eIF4G1, eIF4E, eIF4A1 and eIF4E-BP2 in CD4 + T cells are shown as numbers. (H) Protein copy numbers for eIF2S1, eIF2S2 and eIF2S3 in CD4 + T cells are shown as numbers. The data represented is the average value of a pool of three mice (n=3 pools). Relative protein abundance is graded from low (blue) to high (red). All the data are shown as mean ± SEM. In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤ 0.01.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Mass Spectrometry, Quantitative Proteomics

Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: Changes in proteins involved in DNA replication and cell cycle during CAC CD4 + T cells were sorted from mLN of AOM/DSS-treated mice at days 0 (Healthy), 15 (Colitis) and 60 (Cancer) and their proteomes analysed by quantitative mass spectrometry. (A-B) Expression profiles of key components involved in (A) DNA replication and (B) cell-cycle and transcriptional processes during CAC stages. Data are represented as protein copy numbers per cell (Molecules (n°)) in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). All the data are shown as mean ± SEM. In the centre of panel (A) is a schematic image of DNA replication fork and its associated processing machinery (generated with Biorender). In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Mass Spectrometry, Expressing, Generated

Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: Regulation of glucose metabolism of T cells during CAC Mice were treated with AOM/DSS, and at days 0 (Healthy), 15 (Colitis) and 60 (Cancer), CD4 + T cells were sorted from mLN and their proteomes analysed by quantitative mass spectrometry. (A) Schematic representation of anaerobic glycolysis. Image created with BioRender. (B) Total glycolytic enzymes mass per cell in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (C) Heat map of proteins involved in glycolysis (GO:0006096) identified in CD4 + T cells proteomes in healthy, colitis, and cancer mice. BR, biological replicate. Relative protein abundance is graded from low (blue) to high (red) to allow comparisons. (D) Expression profiles of glycolytic enzymes during CAC progression. Data are represented as protein copy numbers per cell (Molecules (n°)) in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (E-F) Abundances of (E) the glucose transporters Scl2a1 and Slc2a3 and (F) the lactate transporter Slc16a1 in CD4 + T cells. Data are represented as protein copy numbers per cell (Molecules (n°)). Each dot represents a pool of three mice (n=3 pools). (G) Real-time monitorization of extracellular acidification rate (ECAR) in CD4 + T cells along CAC progression. Arrow indicates the time of injection of glucose. (H) ECAR profile of CD4 + T cells in basal state in healthy, colitis, and CAC (n= 4-5 mice). (I) ECAR profile of CD4 + T cells before (-) and after (+) glucose treatment (n= 4-5 mice per condition). All the data are shown as mean ± SEM. In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Mass Spectrometry, Quantitative Proteomics, Expressing, Injection

Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: CD4 + and CD8 + T cells were sorted from mLN at days 0 (Healthy), 15 (Colitis) and 60 (Cancer) from AOM/DSS-treated mice and their proteomes analysed by quantitative mass spectrometry. (A) Heat map of proteins involved in the tricarboxylic acid (TCA) cycle identified in CD4 + and CD8 + T cells proteomes during CAC progression. BR, biological replicate. Relative protein abundance is graded from low (blue) to high (red) to allow comparisons. (B) Expression profiles of key enzymes involved in the TCA/Krebs cycle in T cells in healthy, colitis, and cancer mice. Data are represented as protein copy numbers per cell (Molecules (n°)) in CD4 + and CD8 + T cells. Each dot represents a pool of three mice (n=3 pools). (C-D) (C) Total mitochondria mass and (D) Total OxPhos-related protein mass per cell in CD4 + (left) and CD8 + (right) T cells. Each dot represents a pool of three mice (n=3 pools). (E) Heat map of proteins involved in oxidative phosphorylation process (GO:0006119) identified in CD4 + T cells proteomes in healthy, colitis, and cancer. BR, biological replicate. Relative protein abundance is graded from low (blue) to high (red) to allow comparisons. (F) Protein copy numbers per cell (Molecules (n°)) of the ATP synthase proteins Atp5f1a, Atp5f1b, Atp5f1c, Atp5f1d and Atp5f1e in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (G) Real-time monitorization of oxygen consumption rate (OCR) in CD4 + (left) and CD8 + (right) T cells (n= 3-5 mice per condition). Arrows indicate the time of injection of oligomycin, FCCP and rotenone/antimycin A. (H) OCR profile of CD4 + (left) and CD8 + (right) T cells in basal state in healthy, colitis, and CAC (n= 3-5 mice per condition). (I) Spare respiratory capacity (SRC) profile of CD4 + (left) and CD8 + (right) T cells in healthy, colitis and cancer (n= 3-5 mice per condition). All the data are shown as mean ± SEM. In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Mass Spectrometry, Quantitative Proteomics, Expressing, Phospho-proteomics, Injection

Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: Mice were treated with AOM/DSS, and at days 0 (Healthy), 15 (Colitis) and 60 (Cancer), CD4 + T cells were sorted from mLN and their proteomes analysed by quantitative mass spectrometry. (A) Volcano plot showing the differential expression of proteins associated with T cell activation (GO:0042110) and DNA-binding transcription factor activity (GO:0003700) in Cancer versus Healthy CD4+ T cells. The horizontal lines indicate a p -value of 0.05. The vertical lines indicate a fold change of ± 1.5. (B) Expression profiles of transcription factors related to T cell activation during CAC progression. Data are represented as protein copy numbers per cell (Molecules (n°)) in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (C) Heat map of proteins associated with the “positive regulation of cytokine production” (GO:0001819) identified in CD4 + T cells proteomes during CAC progression. BR, biological replicate. Relative protein abundance is graded from low (blue) to high (red) to allow comparisons. (D) Relative mRNA expression of the different genes at the indicated CAC stages in CD4 + T cells. Actb was used as control. Each dot represents a single mouse (n= 4). R.U.= relative units. (E) Copy numbers per cell (Molecules (n°)) of CD4 + Treg-associated markers FOXP3, ICOS, STAT5, CTLA4, and PD-1. Each dot represents a pool of three mice (n=3 pools). (F) Representative flow cytometry plots (Healthy, Colitis and Cancer) and percentages of regulatory CD4 + T cells (CD45 + CD4 + Foxp3 + ) in mLN from healthy, colitic and cancer-bearing mice. Each dot represents a single mouse (n=3-5). All the data are shown as mean ± SEM. In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Mass Spectrometry, Quantitative Proteomics, Activation Assay, Binding Assay, Activity Assay, Expressing, Control, Flow Cytometry

Journal: bioRxiv

Article Title: Differential proteomic and metabolic remodelling of CD4⁺ and CD8⁺ T cells in mesenteric lymph nodes during colitis-associated colorectal cancer

doi: 10.64898/2025.12.26.696593

Figure Lengend Snippet: Mice were treated with AOM/DSS, and at days 0 (Healthy), 15 (Colitis) and 60 (Cancer), CD4 + and CD8 + T cells were sorted from mLN. (A-B) Volcano plots showing the difference in the expression of proteins between Cancer and Colitis in (A) CD4 + T cells and (B) CD8 + T cells. The horizontal lines indicate a p -value of 0.05. The vertical lines indicate a fold change of ± 1.5. The percentage and number of proteins that are downregulated (blue), upregulated (red), or unchanged (grey) are indicated. (C) Bubble plot showing the enrichment analysis of GO biological processes of the differentially upregulated proteins between Cancer and Colitis in CD4 + T cells. Bubble size indicates the number of genes involved in each process, and bubble color reflects the fold-change between Cancer and Colitis. (D) Left: Heat map of proteins involved in the exocytosis process (GO:0006887) identified in CD4 + T cells proteomes during CAC progression. BR, biological replicate. Relative protein abundance is graded from low (blue) to high (red) to allow comparisons. Right: Copy numbers per cell (Molecules (n°)) of the proteins associated with the exocytosis process STXBP5, RAB11B, SNAPIN, PFN2, PAK1, RAB8A and MYOG1 in CD4 + T cells. Each dot represents a pool of three mice (n=3 pools). (E) Heatmap of proteins and protein copy numbers involved in Methyltransferase activity (GO:0008168) identified in CD4 + T cells in AOM/DSS-treated mice. Relative protein abundance is graded from low (blue) to high (red) per row. BR, biological replicate. Protein copy numbers per cell (Molecules (n°)) of a selection of molecules are shown. Each dot represents a pool of three mice (n=3 pools). All the data are shown as mean ± SEM. In all panels, ns = non-significant, * p ≤ 0.05, ** p ≤0.01, *** p ≤ 0.001.

Article Snippet: On day 6 after AOM injection, mice underwent a 5-day treatment with 2% DSS (w/v) (MW 36,000-50,000 Da; MP Biomedicals, 160110) in drinking water, followed by 15 days administration of normal drinking water.

Techniques: Expressing, Quantitative Proteomics, Activity Assay, Selection