Panel A shows U M A P plots of O T I C D 8 positive T cells clustered into T p e x T e x and cycling populations within tumor microenvironment. Panel B shows U M A P plots of marker gene expression including T c f 7 C c r 7 S l a m f 6 T o x P d c d 1 H a v c r 2 and C x c r 6 across clusters. Panel C shows bubble plots of gene expression patterns highlighting distribution of marker genes across T p e x T e x and cycling clusters. Panel D shows volcano plots of M e t t l gene expression differences comparing T p e x and T e x cell populations. Panel E shows bubble plots of M e t t l 8 expression distribution across different T cell clusters. Panel F shows U M A P plots of M e t t l 8 expression intensity across clustered T cell populations. Panel G shows bubble plots of exhaustion and progenitor markers before and after anti P D 1 treatment. Panel H shows bubble plots of M e t t l 8 expression changes before and after anti P D 1 treatment. Panel I shows box plots of M e t t l 8 expression in tumor infiltrating T cells comparing responders and non responders across cancer types. Panel J shows schematic diagram of tumor implantation and analysis timeline in M e t t l 8 reporter mice. Panel K shows flow cytometry plots and histograms of M e t t l 8 expression in T p e x and T e x cell subsets. Panel L shows schematic diagram of adoptive transfer tumor model using labeled O T I T cells. Panel M shows flow cytometry plots and graphs of M e t t l 8 expression across different T cell division stages.
Mettl8 is highly expressed in T PEX cells and decreased after anti–PD-1 treatment. (A) UMAP plot showing the clustering of OT-I cells infiltrating the MC38-OVA tumor. (B and C) UMAP (B) and bubble (C) plots showing the marker genes expression in each cluster. (D) Volcano plot displaying the fold change expression of Mettl genes in TPEX cells compared with TEX cells. Genes were considered significant if the adjusted P value (FDR) was <0.01. (E and F) Bubble (E) and UMAP (F) plots showing the expression of Mettl8 in each cluster. (G and H) Bubble plots showing the expression of HAVCR2, TOX, TCF7 (G), and METTL8 (H) in the CD8+ T cells of cancer patients before (treatment-naïve) and after (response) anti–PD-1 treatment. (I) Relative expression of METTL8 in tumor-infiltrating T cells from anti–PD-1/CTLA-4–treated melanoma (left) and anti-CD19 CAR-T–treated chronic lymphocytic leukemia (CLL) (right) responders (R) and nonresponders (NR) and in TILs from nivolumab treated NSCLC (middle) R and NR assessed by Tres (https://resilience.ccr.cancer.gov/). Each dot represents a tumor with the average value among all cells on the y axis. The thick line represents the median value. The bottom and top of the boxes are the min and max, respectively. (Melanoma: R = 9, NR = 10; NSCLC: R = 4, NR = 12; CLL: R = 3, NR = 11.) (J) Schematic diagram of the tumor model: Mettl8-tdTomato-Flag mice were subcutaneously injected with 2 × 105 B16F10 cells. Mice were harvested at 13 dpi. (K) Representative cytometry plots showing the Ly108+ Tim3− TPEX and Tim3+ Ly108− TEX cells (left). Cells are gated on tumor-infiltrating CD8+ CD44+ T cells. Representative cytometry plots (middle) and statistic diagram (right) showing the expression of Mettl8-tdTomato in these cells. n = 6–7 per group. (L) Schematic diagram of the tumor model: CD45.1 mice were subcutaneously injected with 2 × 105 EG7-OVA cells, followed by 2 × 106 CTV-labeled Mettl8-tdTomato-Flag OT-I cells transfer at 9 dpi. Mice were harvested at 12 dpi. (M) Representative cytometry plots showing the divisions of CD44+ OT-I cells at day 3 after transfer (left). Representative cytometry plots (middle) and statistic diagram (right) showing the expression of Mettl8-tdTomato in each division. The mean fluorescence intensity (MFI) of tdTomato in division 1, 2, and 3 are all compared with that in division 0. n = 5 per group. Data are representative of two independent experiments. P values were calculated two-tailed Student’s t test, **P < 0.01; ****P < 0.0001.