Intracellular trafficking of anthrax toxins

Anthrax toxins are the major virulence factor secreted by Bacillus anthracis during anthrax pathogenesis. They are produced as three independent multi-domain proteins which associate on the cell surface and subsequently enter the cell hijacking the endocytic route. Edema factor (EF) and lethal factor (LF) are two enzymatic subunits, whose action results in the alteration of two important cellular signaling pathways. They bind to cell surface receptors via interaction with the protective antigen (PA) binding subunit. PA is also essential for EF and LF exit from the endosomal route and delivery to the cell cytoplasm, where they act on their cellular targets. Owing to their enzymatic activity in the host cells cytosol, anthrax toxins play a key role during anthrax pathogenesis, and therefore the knowledge on EF and LF intracellular trafficking is highly important. By imaging EF and LF C-terminally fused to fluorescent proteins in single cell we report that EF and LF reach late endosomal compartments wherefrom they translocate into the cell cytosol. We also show that after cytoplasmic translocation EF and LF have different localization, with LF dispersed into the cytosol and EF associated to endosomal membranes. The interaction between EF and late endosomal membranes is also confirmed in vitro by Surface Plasmon Resonance. By using a domain swap approach we excluded the involvement of EF N-terminal domain in such interaction. Given the importance of compartmentalization of cellular signaling, we point out late endosomes as the encounter platform between anthrax toxins and their targets, highlighting compartmentalization of toxin activity as a key feature in their mechanism of action.

Le tossine di antrace rappresentano il maggior fattore di virulenza secreto da Bacillus anthracis durante la patogenesi dell'antrace. Sono prodotte come tre proteine indipendenti che si associano sulla superficie cellulare e successivamente vengono internalizzate dalle cellule ospite sfruttando la via endocitica. Edema factor (EF) e lethal factor (LF) sono due subunità enzimatiche. Le tossine di antrace hanno un ruolo chiave durante la patogenesi, dunque lo studio del trafficking intracellulare di EF ed LF è molto importante. Questo studio si propone di studiare il trafficking intracellulare d i EF ed LF fuse a proteine fluorescenti in singola cellula e di analizzarne la localizzazione citoplasmatica in cellule target.