AN INDUCED PROXIMITY MODEL FOR CASPASE-8 ACTIVATION

The assembly of the CD-95 (Fas/Apo-1) receptor death-inducing signalin g complex occurs in a hierarchical manner; the death domain of CD-95 b inds to the corresponding domain in the adapter molecule Fas-associate d death domain (FADD) Mort-l, which in turn recruits the zymogen form of the death protease caspase-8 (FLICE/Mach-1) by a hemophilic interac tion involving the death effector domains, Immediately after recruitme nt, the single polypeptide FLICE zymogen is proteolytically processed to the active dimeric species composed of large and small catalytic su bunits, Since all caspases cleave their substrates after Asp residues and are themselves processed from the single-chain zymogen to the two- chain active enzyme by cleavage at internal Asp residues, it follows t hat an upstream caspase can process a downstream zymogen. However, sin ce FLICE represents the most apical caspase in the Fas pathway, its mo de of activation has been enigmatic, We hypothesized that the FLICE zy mogen possesses intrinsic enzymatic activity such that when approximat ed, it autoprocesses to the active protease. Support for this was prov ided by (i) the synthesis of chimeric F(pk)3FLICE molecules that can b e oligomerized in vivo by the synthetic cell-permeable dimerizer FK101 2H2, Cells transfected with F(pk)3FLICE underwent apoptosis after expo sure to FR1012H2; (ii) the creation of a nonprocessable zymogen form o f FLICE that retained low but detectable protease activity.