On the toxic proteins abrin and ricin. Studies of their binding to and entry into Ehrlich ascites cells.

The mechanism of penetration of the toxic plant proteins abrin and ricin into Ehrlich ascites cells has been studied by measuring, under different experimental conditions, the lag occurring from the time the cells have been incubated with the toxins until protein synthesis in the cells is inhibited. When Ehrllch ascites cells were incubated with abrln or ricin at 37”, a lag time of at least 30 min occurred before protein synthesis, as measured by the incorporation of labeled leucine into acid-precipitable material, was reduced. At 20” the lag time was 6 hr. In a cell-free system from rabbit reticulocytes, the inhibition occurred immediately, indicating that the lag time observed in whole cells is due to slow penetration of the toxic principle into the cells. When antisera against the toxins were added to the cell suspension after preincubation with the foxins, the toxic effects could be prevented for different lengths of tie, depending on the incubation temperature and the nature of the antitoxin used. With antisera prepared against the whole toxins, the toxic effects were partially prevented when the antitoxins were added after less than 30 min of preincubation with the cells at 20”. Specific antisera against the isolated A chains (the toxic principle of the toxin) were able to rescue the cells after a longer preincubation with toxins than specific antisera against the B chains which are involved in the binding of the toxins to the cells. Abrin and rlcin did not inhibit protein synthesis in reticulocytes, even though the toxins do hind readily to the surface membrane of these cells. The results are consistent with our two-step model for the entry of abrin and ricin into the cells; in the first step of this model the toxins bind to receptors on the cell surface by their B chain (here denoted the hapLomer), whereas in the second step the toxic A chain (here denoted the efecfomer) penetrates into the cytoplasm, where it exerts its action. The latter process appears to be the slower one, and the data indicate that endocytosis may be involved in this process.