Host cell perforation by listeriolysin O (LLO) activates a Ca-dependent cPKC/Rac1/Arp2/3 signaling pathway that promotes internalization independently of membrane resealing.

Host cell invasion is an indispensable step for a successful infection by intracellular pathogens. Recent studies identified pathogen-induced host cell plasma membrane perforation as a novel mechanism used by diverse pathogens (, , and adenovirus) to promote their internalization into target cells. It was concluded that and adenovirus damage the host cell plasma membrane to hijack the endocytic-dependent membrane resealing machinery, thereby invading the host cell. We studied and its secreted pore-forming toxin listeriolysin O (LLO) to identify key signaling events activated upon plasma membrane perforation that lead to bacterial internalization. Using various approaches, including fluorescence resonance energy transfer imaging, we found that the influx of extracellular Ca subsequent to LLO-mediated plasma membrane perforation is required for the activation of a conventional protein kinase C (cPKC). cPKC is positioned upstream of Rac1 and the Arp2/3 complex, which activation leads to F-actin--dependent bacterial internalization. Inhibition of this pathway did not prevent membrane resealing, revealing that perforation-dependent endocytosis is distinct from the resealing machinery. These studies identified the LLO-dependent endocytic pathway of and support a novel model for pathogen uptake promoted by plasma membrane injury that is independent of membrane resealing.