(A) Frequencies of cytotoxic (granzyme B⁺perforin⁺) CXCR5⁺CD8⁺ T cells in individuals with acute (TopHIVFUTURE: n = 27; circles) and chronic (n = 10; triangles) HIV infection prior and during ART as well as in individuals naturally controlling HIV (LTNPs; n = 20; diamonds) ex vivo. P values by Kruskal-Wallis test with Dunn’s correction for multiple comparisons. (B) Correlation between HIV-specific CD107a expression on CXCR5⁺CD8⁺ T cells responding to overnight stimulation with HIV gag peptide pool at baseline and intact proviral HIV DNA after 48 weeks of ART in individuals treated during acute infection (TopHIVFUTURE; n = 25). Spearman’s rank correlation was performed. (C) Heatmap of genes significantly differentially regulated in individuals treated during acute HIV infection (n = 4; week 4 [w4] versus week 48 [w48] after ART initiation) in HIV-specific CXCR5⁺ and CXCR5^– cells as well as tetramer-negative CXCR5⁺CD8⁺ T cells categorized in functional groups. Relative changes in gene expression between w4 and w48 on ART depicted for each cell type. Normalized read counts, calculated with DESeq2, were converted into z scores and mean z scores per gene significantly differentially expressed across the time points are plotted. Plotted DEGs were selected based on their function. Red indicates higher and blue lower expression. (D) Enriched GO terms (biological process) for significantly downregulated (left) and upregulated (right) genes in HIV-specific CXCR5⁺CD8⁺ and CXCR5^–CD8⁺ T cells w4 versus w48 after ART initiation (BH-adjusted P values < 0.01). The line length represents the ratio of the number of observed DEGs per pathway to the total number of DEGs, and dot size represents the absolute observed number of DEGs in CXCR5⁺CD8⁺ versus CXCR5^–CD8⁺ T cells per pathway. Line and dot color code is according to the adjusted P value.