B cells constitute a major component of infiltrating immune cells in colorectal cancer (CRC). However, the characteristics of B cells and their clinical significance remain unclear. In this study, using single-cell RNA sequencing and multicolour immunofluorescence staining experiments, we identified five distinct subtypes of B cells with their marker genes, distribution patterns and functional properties in the CRC tumour microenvironment. Meanwhile, we found a higher proportion of IgG plasma cells in tumour sites than that in adjacent normal mucosal tissues. In addition, the CXCL13-producing CD8 + T cells in the tumour tissues could promote the formation of tertiary lymphoid structure (TLS) B cells, and the CCL28-CCR10 axis is pivotal for IgG plasma cell migration from the periphery of TLSs to the tumour stroma. Finally, we identified four distinct colon immune classes (CICs: A-D) and found that CD20 + B cells within TLSs were enriched in one immune-inflamed or hot tumour group (CIC D). This B cell-rich group, which was characterized by strong antigen presentation, IgG plasma cells accumulation, microsatellite instability-high (MSI-H) and high tumour mutation Abbreviations: CICs, colon immune classes; CO 2 , carbon dioxide; CRC, colorectal cancer; dMMR, DNA mismatch-repair-deficient; GEO, Gene Expression Omnibus; GSVA, gene set variation analysis; HPFs, high power fields; ICB, immune checkpoint blockade; LPS, lipopolysaccharide; MSI-H, microsatellite instability-high; SCENIC, single-cell regulatory network inference and clustering; ssGSEA, single sample gene set enrichment analysis; TCGA-COAD, the Cancer Genome Atlas Colon Adenocarcinoma; TLS, tertiary lymphoid structure; TMB-H, high tumour mutation burden; TME, tumour microenvironment. Jie Xia, Zhangjuan Xie and Gengming Niu contributed equally to this study.
