Fig. 1: Antigen-specific CD4+ T cells shape macrophage gene expression and drive Slamf1 expression on infected cells.

A Graphical representation of experimental design. B PCA plot of RNAseq expression based on 12000 genes with the greatest variance. C–F Volcano plots of Gene set 1 (C), Gene set 2 (D), Gene set 3 (E), and Gene set 4 (F). Data points correspond to all of the DE genes with fold-change > 2 and FDR ≤ 0.1. P values are adjusted for multiple testing following the Benjamini-Hochberg procedure. (C–E) A subset of the top fifteen most highly upregulated genes are shown in red. A subset of IFN-regulated genes is shown in purple. F Selected genes are shown in red. G SLAMF1 normalized read counts from infected, bystander, and uninfected macrophages co-cultured with or without T cells, as indicated. (UI) Uninfected; (Inf) Infected; (Bys) Bystander. Data are presented as mean values ± SD. Each data point is a biological replicate. P values from Brown Forsythe and Welch ANOVA test with Dunnett T3 multiple comparison test. H Heat map of 25 genes that were significantly DE (FDR < 0.1) in the generalized linear model with ‘T cell x infection status’ interaction term and also upregulated by Mtb infection and CD4+ T cells. log2FC for the respective comparisons (A-D) is indicated. Empty, white boxes indicate no significant difference. I, J Slamf1 expression values in BAL macrophages from scRNAseq scaled gene expression. Data is represented as heat maps, with each line representing expression from a single macrophage. BAL were obtained from NHPs that were unvaccinated (Unvac) or vaccinated with BCG by aerosol (AE), high dose intradermal (IDhigh), low dose intradermal (IDlow), or intravenous routes (IV) as described⁴². BALs were obtained 13 weeks (I) and 25 weeks post-vaccination (J) and either unstimulated or stimulated with PPD prior to scRNAseq, as indicated. FDR comparing the IV vaccinated group to all other groups combined is shown. ns- not significant. A Created in BioRender. Philips, J. (https://BioRender.com/x9c1725). C–F Elements created in BioRender. Philips, J. (https://BioRender.com/we24srw). Source data are provided as a Source Data file.