Mycobacterial surface shedding drives bystander cells response during early intracellular infection

Mycobacteria possess a complex cell envelope whose outermost layer plays an underexplored role in host-pathogen interactions. Using Dictyostelium discoideum as a model host phagocyte, we show that Mycobacterium marinum rapidly sheds its envelope components, including surface proteins, carbohydrates, and virulence-associated lipids, within minutes of uptake. Shed material is actively trafficking within host endocytic pathways and disseminates to neighboring bystander cells, where it accumulates and triggers several responses. Notably, bystander cells exposed to shed material exhibit a transient delay in G1/S cell cycle progression, an activation of membrane damage-response pathways, and an enhanced resistance to subsequent mycobacterial infection. These phenotypes are recapitulated by infection-free conditioning with purified envelope extracts, demonstrating that superficial components of the envelope alone are sufficient to modulate host cell responses. Moreover, this priming effect is independent of bacterial viability or the Esx-1 secretion systems, underscoring the intrinsic immunomodulatory capacity of the envelope. Interestingly, bacteria that lose their outer layer are more frequently ubiquitinated, suggesting that host-driven stripping exposes molecules that are recognized by cytosolic sensors to mount a cell-autonomous defense. Together, our findings reveal that mycobacterial envelope shedding is a widespread, early event during intracellular infection that impacts both infected and bystander cells. These findings suggest that mycobacterial outermost envelope components can influence host cell physiology and contribute to early innate immune modulation, with implications for understanding the initial determinants of infection outcomes.