Figure S3.
Two Distinct CD4+ T Cell Subpopulations Are Associated with COVID-19 Severity, Related to Figure 3
A,B. UMAP embedding of all CD4+ T cells colored by unsupervised clustering (top left of A) and by selected mRNA transcript levels (other panels in A) or two selected surface proteins (B).
C. UMAP embedding of all CD4+ T cells colored by the density of cells characterized by different clonal expansion sizes (n = 1, n = 2-4, and n > = 5).
D. Clonal expansion sizes of each CD4+ T cell subset from unsupervised clustering. Bar plot shows the normalized clonal composition.
E. Boxplots showing percentages of naive CD4+ T cells (cluster 1 and 2), cluser5 and 8 over all CD4+ T cells in samples grouped by WOS.
F. Boxplots showing the mRNA expression levels in samples grouped by WOS.
G. Clonal expansion status, presented as bar plots, for each CD4+ T cell cluster, color-coded by clonal expansion sizes (n = 1, n = 2-4, n > = 5). The pie charts show the CD4+ T cell cluster composition for each clonal expansion sizes present in the bar plots
H. Bar plots representing four functional signature scores across different subclusters of CD4+ T cells. P values are shown.
I,J. Flow cytometry validation of cytotoxic CD4+ T cells (I) and proliferative-exhausted cluster 8 CD4+ T cells (J). Left panel: Representative flow cytometry plots of the gating strategy for cytotoxic CD4+ T cells (I), and proliferative-exhausted CD4+ T cells (J). Gating was performed on all CD4+ T cells. Middle panel: quantitative comparison of the % of gated cytotoxic (proliferative-exhausted) CD4+ T cells (y axis) and cluster 5 (cluster 8) CD4+ T cell % (x axis); each dot represents a PBMC sample. The regression line is drawn in orange. Pearson correlation coefficient and associated P value shown. Right panels: Boxplots represent % of cytotoxic (proliferative-exhausted) CD4+ T cells quantified by flow cytometry and % of cluster 5 (cluster 8) cells quantified by 10X single cell RNA-seq. Comparison was between samples from healthy donor and COVID-19 patients. Data are represented as mean ± SEM.
K, L. Bar plots represent GSEA of top pathways enriched for cluster 5 (K) and cluster 8 (L) cells. Normalized enrichment score (NES) and P values are shown. Full results provided in Tables S3.1 and S3.2.
M. Heatmap visualization of average cytokine secretion frequencies for cells from healthy donor (HD), mild (WOS = 1-2), moderate (WOS = 3-4) and severe (WOS = 5-7) patient samples.
Significance is indicated by: (∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001).