Ribosome-inactivating toxins such as ricin and Shiga-like toxins have been used in the design of cancer therapeutics1.  Knowing how these toxins are taken-up and processed by eukaryotic cells represent an important source of information in designing second-generation targeting agents.  Shiga-like toxin 1 (SLT-1) exerts its cytotoxic effect by retrotranslocating its toxic domain to the cytosol from the ER lumen (Figure 1 and Figure 2)² It is the current belief that existing ER mechanisms are exploited by ER-routing toxins such as SLT-1 to escape the ER lumen and cause toxicity. Our research focuses on the use of chemical and genetic approaches to define the mechanism that the A1 chain of SLT-1 exploits, to escape the ER lumen.

Figure 1: Mapping the minimal domain of the A chain of Shiga-like toxin 1 responsible for its catalytic property and its ability to retrotranslocate outside the ER lumen ¹.

Figure 2: Delimiting the miniumum cytotoxic domain of SLT-1 and domain of ER escape.
(A) 10 fold serial dilutions of yeast expressing a catalyticallyinactive mutant (DETOX), full length A1 chain (1-251), or truncated forms (1-239, 1-238,6-251, 7-251, 8-251) spotted on SD agar -URA plus 2% galactose to activate the expression of the protein. Toxicity is ablated with the deletion of residues 1-7 and 239-251.
(B) 10 fold serial dilutions of yeast expressing ER- targeted C-terminal truncations of the A1 chain.The C-terminal truncation of ER expressed toxin variants shows that residues 239-251 are required for retrotranslocation into the cytosol from the ER lumen ².

References:

1. Bray MR, Bisland S, Perampalam S, Lim WM, Gariépy J. (2001). Probing the surface of eukaryotic cells using combinatorial toxin libraries. Curr Biol. 11(9):697-701.
   
   
2. LaPointe, P, Wei X, and Gariépy J. (2005). A Role for the Protease-sensitive Loop Region of Shiga-like Toxin 1 in the Retrotranslocation of Its A1 Domain from the Endoplasmic Reticulum Lumen. J. Biol. Chem. 280;23310-23318.


3. McCluskey AJ, Poon GMK, Bolewska-Pedyczak E, Srikumar T, Jeram, SM, Raught B, Gariepy J. (2008). The catalytic subunit of shiga-like toxin 1 interacts with ribosomal stalk proteins and is inhibited by their conserved C-terminal domain. J Mol Biol. J. Biol. Chem. 378(2):375-86.