Temporal single-cell RNA sequencing captures macrophage reprogramming by Salmonella

While macrophages are well-equipped to track and kill foreign invaders, they paradoxically constitute a replicative niche for several intracellular pathogens. For instance, the intracellular Gram-negative bacteria Salmonella enterica serovar Typhimurium can turn infected macrophages into a favourable intracellular niche by secreting bacterial proteins effectors within the host cytoplasm. Typically, macrophages induce a pro-inflammatory program (coined M1) upon bacterial invasion to control and kill the bacteria. However, Salmonella can reverse macrophage polarization towards an anti-inflammatory state (so-called M2) permissive to bacterial survival and replication. The understanding of the transcriptional cascade leading to this reprogramming on the macrophage side remains unknown. Here, we leveraged temporal single-cell RNA-sequencing and RNA metabolic labelling to resolve the transcriptomic response of mouse bone marrow-derived macrophages during the first hours following Salmonella challenge.

The initial and stereotypical response driving infected macrophages to M1 polarization was characterized by the pathogen detection and the strong induction of NF-κB signaling. Heterogeneity emerged in the macrophage population at 6 hours following infection, with some infected cells switching to the M2 phenotype. This critical point determined the final polarization state of macrophages. The M2 macrophage population was marked by a clear dampening of the initial inflammatory response and the induction of specific gene modules, concurrently to the activation of the Salmonella pathogenicity island 2 (SPI2). We identified transcription factors marking the initiation of the reprogramming trajectory and correlated to the M2 polarization even at later timepoints. Overall, our work highlights a weak point in macrophage wiring exploited by a pathogen to overcome cellular immunity and induce host cell reprogramming.