The pore-forming toxin proaerolysin is activated by furin

Aerolysin is secreted as an inactive dimeric precursor by the bacterium Aer omonas hydrophila. Proteolytic cleavage within a mobile loop near the C ter minus of the protoxin is required for oligomerization and channel formation . This loop contains the sequence KVRRAR(432), which should be recognized b y mammalian proprotein convertases such as furin, PACE4, and PC5/6A. Here w e show that these three proteases cleave proaerolysin after Arg-432 in vitr o, yielding active toxin. We also investigated the potential role of these enzymes in the in vivo activation of the protoxin. We found that Chinese ha mster ovary cells were able to convert the protoxin to aerolysin in the abs ence of exogenous proteases and that activation did not require internaliza tion of the toxin, The furin inhibitor alpha(1)-antitrypsin Portland reduce d the rate of proaerolysin activation in vivo, and proaerolysin processing was even further reduced in furin-deficient FD11 Chinese hamster ovary cell s, The cells were also less sensitive to proaerolysin than wild type cells; however, transient transfection of FD11 cells with the cDNA encoding furin conferred normal sensitivity to the protoxin. Together these findings argu e that furin catalyzes the cell-surface activation of proaerolysin in vivo.