FIGURE 6.

Ser-62 to Ala mutation of Bcl-xL renders cells highly resistant to vinblastine-induced apoptosis. A, cells expressing wild-type HA-Bcl-xL or two clones expressing the S62A mutant or one clone expressing the T47A mutant were untreated or treated with vinblastine (200 nm, 48 h), and the relative extent of apoptosis was assessed quantitatively using a cell death enzyme-linked immunosorbent assay as described under “Experimental Procedures.” Values on the y axis represent absorbance at 405 nm. The results represent the mean ± S.D. (n = 6) and are representative of two independent experiments. *, significantly different from cells expressing WT Bcl-xL (p < 0.001). B, cells expressing wild-type HA-Bcl-xL or the S62A mutant were untreated or vinblastine-treated (200 nm) for 36 h, and cell extracts were prepared and subjected to immunoblotting for poly(ADP-ribose) polymerase (PARP). The blots were also probed for Bcl-xL and GAPDH expression as indicated. C, cells expressing wild-type HA-Bcl-xL or the S62A mutant were untreated or vinblastine-treated (200 nm) for 36 h, and cell extracts were prepared and subjected to immunoprecipitation with 6A7 active Bax antibody and immunoblotting for Bax. A precipitate prepared in the absence of 6A7 antibody was analyzed as control. D, whole cell extracts were immunoblotted for total Bax expression and reblotted for GAPDH to confirm equal loading.