EXPRESSION OF MUTANT DYNAMIN PROTECTS CELLS AGAINST DIPHTHERIA-TOXIN BUT NOT AGAINST RICIN

Diphtheria toxin is believed to enter sensitive mammalian cells via re ceptor-mediated endocytosis from clathrin-coated pits, while ricin can enter via both clathrin-dependent and clathrin-independent endocytosi s. The present study has confirmed this by determining the toxin sensi tivity of COS-7y cells which were transiently overexpressing a trans d ominant negative mutant of dynamin, a GTPase required for the budding of clathrin-coated vesicles from the plasma membrane. Cells overexpres sing wild-type dynamin showed normal receptor-mediated endocytosis of transferrin and remained sensitive to both diphtheria toxin and ricin. Cells overexpressing a mutant dynamin defective in GTP binding and hy drolysis were unable to endocytose transferrin and were protected agai nst diphtheria toxin, but they remained completely sensitive to ricin intoxication. Treating nontransfected cells or cells overexpressing mu tant dynamin with nystatin caused a redistribution of the caveolae mem brane marker protein VIP21-caveolin from the cell surface to intracell ular locations, but did not affect their sensitivity to ricin. The red istribution of caveolin seen after nystatin treatment may reflect the disappearance of caveolae. If this is the case, caveolae are not respo nsible for the endocytosis of ricin. An alternative clathrin-independe nt route may operate for ricin, since cellular uptake, intracellular t ransport, and translocation into the cytosol remain unaffected when cl athrin-dependent endocytosis is effectively blocked. (C) 1998 Academic Press.