R. Wales et al., ADDITION OF AN ENDOPLASMIC-RETICULUM RETRIEVAL SEQUENCE TO RICIN-A CHAIN SIGNIFICANTLY INCREASES ITS CYTOTOXICITY TO MAMMALIAN-CELLS, The Journal of biological chemistry, 268(32), 1993, pp. 23986-23990
An Escherichia coli expression system was used to produce recombinant
ricin A chain (RTA) and RTA modified either by the addition of a carbo
xyl-terminal endoplasmic reticulum retrieval sequence Lys-Asp-Glu-Leu
(RTAKDEL) or a nonfunctional analogue Lys-Asp-Glu-Ala (RTAKDEA). These
RTA molecules can enter mammalian cells by fluid phase endocytosis. R
TAKDEL was significantly more cytotoxic than either RTA or RTAKDEA to
both Vero cells and HeLa cells (250- and 10-fold, respectively), despi
te the fact that all these RTA molecules had comparable enzymatic acti
vities. This difference did not result from KDEL-mediated binding of R
TAKDEL to the cell surface. Enhanced cytotoxicity could be correlated
with an increased level of ribosome inactivation, measured as the RTA-
catalyzed depurination of 28 S ribosomal RNA. These results indicate t
hat the added KDEL sequence facilitated RTA entry into the cytosol. We
propose that interaction with the intracellular KDEL receptor promote
s retrograde transport of the toxin to the endoplasmic reticulum, wher
e translocation of RTA into the cytosol occurs.