Fig 1: EphA4 on T cells may contribute to RNase1-mediated antitumor immunity. (A) Quantitative analysis of RNA-sequencing results from Human Protein Atlas (Monaco database) of EphA4 expression on immune cells. (B) Quantitative analysis of EphA4 gene expression on a single-cell level of human PBMCs based on markers of different cell type lineages. (C) Flow cytometric analysis of EphA4 and CD3 expression in human PBMCs. IgG-APC and secondary Ab only serve as negative controls. Frequencies of cell population after gating are indicated in the quadrant of each panel. (D and E) Time-course quantitation of T cell-meditated tumor cell-killing assay of dead cells, normalized to that at the zero-time point, in BT-549-aCD3 cells cocultured with human PBMCs (D) or primary T cells (E) combined with RNase1 treatment (1 µg/ml) in the presence or absence of recombinant His-A4 (5 µg/ml) as indicated. Data represent mean ± SEM of three independent experiments. ****p < 0.0001, ANOVA test. (F) A proposed model of RNase1-mediated T cell killing towards tumor cells in the breast TME. In brief, serum RNase1 known as a ligand of EphA4 promotes oncogenesis in breast cancer cells. However, T cell function plays a dominate role in the presence of the immune system, namely that RNase1 boosts CD4+ T cell activation, in which EphA4 may participate, leading to activated T cell-mediated antitumor immunity. Artwork, adapted from “T-cell Deactivation vs. Activation”, by BioRender.com (2023). Retrieved from https://app.biorender.com/biorender-templates