Inhibition of the unfolded protein response by ricin a-chain enhances its cytotoxicity in mammalian cells

Ricin is a highly toxic type II ribosome-inactivating protein that has potential as a biochemical weapon and as the toxic component of immunotoxins. The unfolded protein response (UPR) is a survival response that helps cells to recover from endoplasmic reticulum (ER) stress. Failure to recover from ER stress leads to apoptosis. In yeast, ricin-A-chain (RTA), the enzymatic component of ricin, inhibits UPR. Our goals were to determine if RTA inhibits UPR in two epithelial cell lines and if this affects RTA cytotoxicity. RTA alone did not induce UPR. However, RTA inhibited both phosphorylation of inositol-requiring enzyme 1 (IRE1) and splicing of X-box binding protein1 mRNA by the UPR-inducing agent tunicamycin (Tm). The ability of dithiothreitol (DTT) to activate eukaryotic translation initiation factor 2 alpha (eIF2α), a component of the PERK pathway, was also inhibited by RTA. Treatment with RTA in combination with Tm or DTT inhibited protein synthesis more than either agent did alone in one cell line, while caspase cleavage was enhanced by the treatment combination in both cell lines. These data indicate that RTA is more cytotoxic when UPR is inhibited. This ability to inhibit UPR may enhance the potential of RTA as a therapeutic immunotoxin in solid tumors.