Repair of a Bacterial Small β-Barrel Toxin Pore Depends on Channel Width.

Membrane repair emerges as an innate defense protecting target cells against bacterial pore-forming toxins. Here, we report the first paradigm of Ca-dependent repair following attack by a small β-pore-forming toxin, namely, plasmid-encoded phobalysin of subsp. In striking contrast, cytolysin, the closest ortholog of phobalysin, subverted repair. Mutational analysis uncovered a role of channel width in toxicity and repair. Thus, the replacement of serine at phobalysin´s presumed channel narrow point with the bulkier tryptophan, the corresponding residue in cytolysin (W318), modulated Ca influx, lysosomal exocytosis, and membrane repair. And yet, replacing tryptophan (W318) with serine in cytolysin enhanced toxicity. The data reveal divergent strategies evolved by two related small β-pore-forming toxins to manipulate target cells: phobalysin leads to fulminant perturbation of ion concentrations, closely followed by Ca influx-dependent membrane repair. In contrast, cytolysin causes insidious perturbations and escapes control by the cellular wounded membrane repair-like response. Previous studies demonstrated that large transmembrane pores, such as those formed by perforin or bacterial toxins of the cholesterol-dependent cytolysin family, trigger rapid, Ca influx-dependent repair mechanisms. In contrast, recovery from attack by the small β-pore-forming alpha-toxin or aerolysin is slow in comparison and does not depend on extracellular Ca To further elucidate the scope of Ca influx-dependent repair and understand its limitations, we compared the cellular responses to phobalysin and cytolysin, two related small β-pore-forming toxins which create membrane pores of slightly different sizes. The data indicate that the channel width of a small β-pore-forming toxin is a critical determinant of both primary toxicity and susceptibility to Ca-dependent repair.