TEMPERATURE-DEPENDENT AND PH-DEPENDENT CYTOTOXIC EFFECT OF THE HEMOLYTIC LECTIN CEL-III FROM THE MARINE INVERTEBRATE CUCUMARIA-ECHINATA ON VARIOUS CELL-LINES

We investigated the cytotoxicity of CEL-III, one of four Ca2+-dependen t galactose/N-acetyl- galactosamine (GalNAc)-binding lectins from the marine invertebrate Cucumaria echinata, Among six cell lines tested, M DCK cells showed the highest susceptibility to CEL-III cytotoxicity an d its LD(50) was estimated to be 53 ng/ml, while no significant cytoto xicity of CEL-III was observed in CHO cells up to 10,000 ng/ml, In the presence of 0.1 M lactose, the cytotoxicity of CEL-III was strongly i nhibited, The binding studies using FITC-labeled GEL-III revealed that the amount of CEL-III bound to MDCK cells was about 2-fold greater th an that in the case of CHO cells, The cytotoxicity of CEL-III increase d with decreasing temperature, The surviving fractions of Vero cells e xposed to CEL-III at 4 degrees C were immediately decreased, and more than 90% of exposed cells were killed within 20 min, whereas at 37 deg rees C much longer exposure period (more than 10 h) was required to ki ll 50% of the cells, CEL-III induced the release of carboxyfluorescein (CF) from CF-loaded MDCK cells and this activity was markedly increas ed at alkaline pH (pH 10) and at lower temperature (4 degrees C), Even in CHO cells, considerable CF release was induced by CEL-III at 4 deg rees C and at pH 10 but not at pH 7.5 at both temperatures, In agreeme nt with these results, CHO cells exposed to CEL-III at 4 degrees C and at pH 10 were killed in a dose-dependent manner, These results sugges t that CEL-III exhibits cytotoxicity through damaging the plasma membr ane by pore-formation in a temperature- and pH-dependent manner, Diffe rent susceptibility of each cell line to CEL-III cytotoxicity may be d ue to differences in the processes leading to pore-formation after bin ding to cell-surface carbohydrates.