Transcriptional profiling identifies 4 distinct memory CD8α⁺ T-cell subsets defined by expression of CD161 and CD62L. (A) Principal component analysis shows grouping of gene expression profiles of T_N (orange), CD161^hi CD62L⁺ (red), CD161^hi CD62L⁻ (green), CD161^lo CD62L⁺ (blue), and CD161^lo CD62L⁻ (purple) subsets isolated from 6 healthy individuals. (B) Hierarchical cluster analysis of gene expression in sort-purified memory subsets compared with matched T_N cells from 6 healthy individuals shows transcriptionally distinct memory subsets. Genes and subsets are clustered in columns and rows, respectively. All samples of each subset clustered together; therefore, only the names of the clustered subsets and not the individual samples are shown in the figure. Red profiles represent higher gene expression (maximum 32-fold) and green profiles represent lower gene expression (minimum 32-fold) compared with matched T_N cells. (C) Volcano plots show differentially expressed transcriptional profiles of CD161^hi CD62L⁺ compared with CD161^lo CD62L⁺ cells (top left), CD161^hi CD62L⁺ compared with T_N cells (top right), CD161^hi CD62L⁻ compared with CD161^lo CD62L⁻ cells (bottom left), and CD161^hi CD62L⁻ compared with T_N cells (bottom right) from 6 healthy individuals. Each point represents a single gene and/or probe, and type 17 associated genes are color-coded, as indicated in the legend. The numerator and denominator defining the mean log₂ ratio of gene expression are listed first and second, respectively, in the volcano plot title; thus, genes expressed at higher levels in the numerator subset are depicted on the right of each plot.