Figure 3. ES_I causes a delay in intoxication of HeLa cells with SLTx.

A, B. The UPS has little influence on SLTx cytotoxicity. A. Cells treated with the proteasome inhibitor clasto Lactacystin β-lactone (cLβ-l) were challenged with SLTx for 1 (left), 2 (middle) or 4 h (right) and remaining levels of protein synthesis were determined. B. Efficacy of cLβ-l was confirmed in vitro by its ability to block degradation of casein (arrowhead) in the presence of mammalian 26S proteasomes. C. Cells treated with ES_R35 (left hand panel) or ES_I (right hand panel) were challenged with a fixed dose (1 µg.ml⁻¹) of SLTx for increasing periods and protein synthesis levels were determined. Exponential fits (dotted lines) were extrapolated to determine the lag phase of intoxication (white arrowhead, DMSO; black arrowhead, drug). Typical single assays are shown. D. Lag phases were determined as in C, and the half-lives of protein synthesis (T_1/2) were determined from exponential fits in C. DMSO, n = 6, +/− 1.S.D., ES_I and ES_R35, n = 3, +/− 1 S.D. Values were compared using unpaired t-tests. Lags: DMSO versus ES_R35, no significant difference; DMSO versus ES_I, p<0.0001, t = 8.347; ES_I versus ES_R35 p = 0.0016, t = 7.658; **, very significant: ***, extremely significant. T_1/2 values: no significant differences.