Transcriptional profile of colonic LP CD4 ⁺ T cells confirms a Th1/Th17 signature and reveals restriction of TCR repertoire. Total CD4⁺ T cells were sorted from the colonic LP of littermate wild-type (+/+) or Rag1^w/w (W/W) mice at 3 and 12 wk of age, and analyzed by scRNA-seq. (A) UMAP plots showing the expression profiles of CD4⁺ T cells split by condition. Colors represent cells clustered together on the basis of similarity of global gene expression. (B) The stacked bar plot shows cluster distribution in each condition. (C) Selected gene ontology (GO) biological process terms enriched among gene sets differentially regulated (with >50% fold change and FDR = 0) in a cell type–specific manner between W/W and +/+ mice at 12 wk of age. The color of each circle represents the FDR value of the enriched GO term, whereas the size of each circle is proportional to the enrichment ratio, which is the number of differentially expressed genes associated with the GO term normalized by the number of such genes that one can encounter by random chance given the size of the total gene set in the GO database. (D) TCR clonotype analysis shows the proportion of different clonotype abundance ranges (single: 1 cell; small: 2–5 cells; medium: 6–20 cells; large: 21–100 cells; and hyperexpanded: 101–500 cells) of total CD4⁺ T cells sorted from the colonic LP of +/+ or W/W at 3 and 12 wk of age and analyzed by scRNA-seq. (E) TCR clonotype analysis projected on UMAP of CD4⁺ T cells split by condition. The color scale matches the abundance range as in D. (F) Hydrophobicity index of the CDR3 region of TRB clonotypes of LP CD4⁺ T cells from +/+ and W/W mice at 3 and 12 wk of gestational age. Single-cell samples were obtained from four to five pooled mice/group. Data are from two independent experiments.