Fig. 5: High degree of heterogeneity in HIV-infected cells from peripheral blood of ART-suppressed PLWH.

a, Differential expression of surface antigens was assessed between all HIV⁺ CD4⁺ T cells (n = 205) versus HIV^– CD4⁺ T cells (n = 146,016). Test performed using the DESeq2 pseudobulk method in Seurat (two-tailed with multiple comparison adjustment using the Bonferroni–Hochberg method). b, Same comparison and cells as in a but using the Wilcoxon statistical test in Seurat (two-sided with multiple comparison adjustment using the Bonferroni method). c, Differential expression of surface antigens was assessed between Tcm/Ttm HIV⁺ (n = 57) versus Tcm/Ttm HIV^– cells (n = 39,954) using the DESeq2 method in Seurat (two-tailed with multiple comparison adjustment using the Bonferroni–Hochberg method). Tcm/Ttm cells are defined by the combination of all clusters containing the terms ‘Tcm’ or ‘Ttm’ or ‘cTfh’ (but not MAIT or recently activated Tcm/Ttm cells) in Fig. 4a. d, Differential expression of surface antigens was assessed between Tem/effector HIV⁺ (n = 59) versus Tem/effector HIV^– (n = 48,928) cells using the DESeq2 method in Seurat (two-tailed with multiple comparison adjustment using the Bonferroni–Hochberg method). Tem cells are defined by the combination of all clusters containing the term ‘Tem’ in Fig. 4a. Markers in a–d are ranked by π-score (see Fig. 1e legend). All markers shown have an adjusted P value < 0.05. e, Comparison of motifs associated with accessible chromatin regions of cells grouped by phenotype (MAIT (n = 58 for HIV⁺; n  =  38,405 for HIV⁻), Tcm/Ttm (n = 85 for HIV⁺; n = 53,135 for HIV⁻; includes recently activated Tcm/Ttm cells) and Tem/effector (n = 59 for HIV⁺; n = 48,928 for HIV⁻)) and infection status (HIV^– versus HIV⁺) as assessed through chromVAR and ArchR. Heatmap displays the mean chromVAR deviations (z score) of transcription factor motifs from CISBP database (displayed by row) that define each aggregate group (displayed by column). Motifs were selected by FDR < 0.05 and a mean difference > 0.5, indicating a significant accessibility of regions that contain a given transcription factor motif for cells in the cluster (indicated by asterisks) as compared with all other cells (getMarkerFeatures function in ArchR). Each cell is colored by the mean deviation (z score) where values greater than zero indicate positive enrichment of a motif. Motifs were clustered using k-means clustering and the motifs are labeled to the right of the heatmap in order from top to bottom for each cluster. Asterisks indicate that the motif was significantly enriched in the specific group (column). *P < 0.05; **P < 0.01. f, Differential chromVAR motif profiles were assessed between HIV⁺ Tcm/Ttm cells and HIV^– Tcm/Ttm cells. The top 20 most significant motifs for HIV⁺ cells are shown. The dotted line indicates a –log₁₀ transformed FDR value of 0.05. Color indicates the mean difference in chromVAR deviations (value greater than zero indicates enrichment in HIV⁺ cells).