Anthrax toxin-mediated delivery of cholera toxin-A subunit into the cytosol of mammalian cells

Abstract

The protective antigen (PA) component of anthrax toxin mediates delivery of either lethal factor (LF) or oedema factor into the cytosol of mammalian cells. The N-terminal domain of LF^1-254 (amino acids 1-254 of LF) binds to PA and, when fused to heterologous proteins, delivers such proteins into the cytosol. In the present study, we fused the catalytic subunit of cholera toxin (CT-A) with LF^1-254 and showed that the fusion protein LF^1-254-CT-A retains ADP-ribosylation activity in solution and increased intracellular cAMP levels in J774A.1 macrophage cells when added together with PA. A mutant fusion protein, in which arginine-7 of CT-A was replaced with lysine, did not show ADP-ribosylation activity in solution and failed to increase cAMP levels in macrophage cells. The data show that LF^1-254-CT-A retains its catalytic activity in solution as well as when translocated into the cytosol of eukaryotic cells via an alternative pathway to the GM₁ receptor used by CT.