FIGURE 2.

Caspase activation and substrate cleavage are concomitant with CD4⁺ T cell activation. Day 4 polyclonal effector CD4⁺ T cells were generated as in Fig. 1. Lysates were prepared from viable cells at the times indicated. Immunoblot analysis was performed for caspase-8 (A), c-FLIP_L (B), Bid (C), caspase-3 (D), and ICAD (E). Caspase-8 represents noncleaved procaspase-8, whereas p43/41caspase-8 bands represent fragments generated by cleavage at Asp³⁷⁴ or Asp³⁸⁴. c-FLIP_L represents full-length protein, p43FLIP represents a fragment generated by cleavage at Asp³⁷⁶, and p27FLIP represents a further cleavage product from cleavage at Asp²⁵⁸. t-Bid is generated by cleavage of full-length Bid at Asp⁷⁵. Caspase-3 represents noncleaved procaspase-3, whereas p20 caspase-3 and p12/10 caspase-3 represent fragments generated by cleavage at Asp¹⁶⁹ or Asp¹⁷⁵. ICAD represents noncleaved ICAD, and p18ICAD is generated by cleavage at Asp¹¹⁷. Lysates from FasL-treated populations were included in the analysis to illustrate cleavage products. NS, Nonspecific band