Sensitivity of Carrot Cell Cultures and RNA Polymerase II to Amatoxins : Evidence for the Inactivation of 6'-Hydroxyamatoxins

Abstract

Protoplast and cell suspension cultures of Daucus carota L. were evaluated for their sensitivity toward the three amatoxin derivatives, alpha-amanitin, 6'-deoxy-alpha-amanitin, and 6'-O-methyl-alpha-amanitin using inhibition of DNA synthesis to measure cell viability. Protoplasts appeared approximately 10-fold more refractory than suspension cells and alpha-amanitin was much less effective than the other two amatoxins, even though K(i) values for isolated RNA polymerase II were similar (4-5 nanomolar). Additional studies evaluating the recoveries of all three amatoxins from cell suspension supernates indicate one basis for these differences to be the selective degradation of alpha-amanitin. A mechanism involving the activation of the hydroxyindole moiety of the alpha-amanitin is thus invoked to explain these differences and we postulate the involvement of plant oxidases in this role.