Tunicamycin on Transformed BALB/3T3 Fibroblasts Selective Cytotoxicity of Purified Homologues of Updated Version

The selective cytotoxicity of tunicamycin homologues against SV40-transformed 3T3 cells (SV40-3T3) was examined. Incubation of 3T3 or virally transformed 3T3 cells with four different homologues (A~, A2, B1, and B2 at 0.1 to 0.25 #g/ml) caused detachment and death of transformed cells after 1 to 3 days, while the nontransformed cells were almost unaffected. Cytotoxicity against nontransformed cells occurred only when higher doses (at least 5-fold) of A2-, B~-, and B2-tunicamycins were used. In contrast, these homologues inhibited proliferation of 3T3 cells, even when doses of 0.5 ffg/ml were used. These cytotoxic effects are dose dependent, and maximal cytotoxicity of each homologue is achieved at a different concentration in each cell type. These results indicate that tunicamycin homologues have selective cytotoxicity against transformed cells. Incorporation of [3H]mannose into acid-precipitable macromolecules synthesized by transformed cells was strongly inhibited (70 to 75%) by AIand B2-tunicamycins at 0.01 to 0.05 ffg/ml, while incorporation by 3T3 cells was not affected. At higher concentrations of the above tunicamycins (0.5 to 1 #g/miX [3H]mannose incorporation by both 3T3 and SV40-3T3 cells was inhibited more than 95%. In contrast, the effect of these tunicamycin homologues on protein synthesis in 3T3 and SV40-3T3 fibroblasts was less pronounced since the incorporation of amino acids was inhibited by approximately 20%. Very little inhibition of amino acid incorporation occurred when 3T3 or SV40-3T3 cells were treated with B2-tunicamycin. However, A~-tunicamycin inhibited [3H]proline incorporation and slightly increased [3H]tyrosine incorporation into cell layers of 3T3 cells. Examination of secreted proteins synthesized by these cells on sodium dodecyl sulfate:polyacrylamide gel electrophoresis revealed that both 3T3 and SV40-3T3 cells treated with homologues produced partially glycosylated macromolecules, such as procollagen and fibronectin, and failed to convert procollagen to collagen. Tunicamycin homologues also inhibited the N-acetylglucosamine-1 -phosphate transferase activity found in microsomes prepared from 3T3 and virally transformed 3T3 fibroblasts. The data presented indicate that the cytotoxic activity of purified homologues of tunicamycin against transformed fibroblasts might be due to the selective inhibition of glycosylation and to the differences in the membrane solubilities of the homologues.