Caspases: Intracellular Signaling by Proteolysis

Ectopic expression of caspase 1 in some cells can result The discovery of a novel proteolytic network in the cytoin apoptosis, but a role in developmentally programmed sol of metazoan cells comes as a surprise to a field that cell death is unlikely given the normal phenotype of previously considered intracellular proteolysis to mainly knockout mice (see for example, Ghayur et al., 1997). be a mechanism to process antigen or remove effete Processing of thecaspase 1 precursor, required for actiproteins. The network, supported by caspases, a family vation, is rarely seen in experimental models of apopof cysteine proteases that specifically cleave proteins tosis, and thus, though it is clearly required for activation after Asp residues, involves limited proteolysis of a of the cytokines mentioned above, the role of this casgrowing numberof cellular substrates. It is not surprising pase in apoptosis, if any, is still uncertain. The close that a signaling pathway utilizes proteolysis, for this is relationship in sequence identity and predicted subthe essence of the blood coagulation cascade. What is strate specificity of caspases 4 and 5 implicate them as surprising is that the caspase pathway takes place in effectors of cytokine activation, possibly as upstream the interior of a cell. Caspase precursors are usually activators of caspase 1 itself. activated at internal conserved Asp residues, with the Whereas caspase 1 (and possibly 4 and 5) is primarily result that most activated caspases can process their involved in procytokine activation, othercaspases, notaown and other caspase zymogens given sufficient time bly 2, 3, 6, 7, 8, and 10, are considered to promote and a high enough concentration in vitro (see for exampathways to apoptosis. This conclusion is based largely ple, Srinivasula et al., 1996). This suggests a cascade on the following observations: (1) thezymogens are seen mechanism for transmission of signals, but the extent to be processed during apoptosis, or in vitro models of to which this happens in vivo is currently an enigma, apoptosis, and (2) at least in vitro, they cut proteins and a fertile area of research. whose cleavage is associated with apoptotic cell death. The caspases seem tobe a developmentof multicelluSo far, about a dozen proteins have been shown to lar organisms, and in humans at least seven of the ten be specifically cleaved during apoptosis. It is relatively currently known family members participate in one of simple to demonstrate which clips are caused by castwo distinct signaling pathways: (1) activation of proinpases (it turns out to be almost all of them) through flammatory cytokines, and (2) promotion of apoptotic analysis of the cleavage site, but more difficult to detercell death. Recent publications have reviewed the dismine which caspases are responsible in vivo. For examcovery of the caspases and some of their substrates ple, the first protein discovered to be cleaved by cas(see for example, Nicholson, 1996; Zhivotovsky et al., pases during apoptosis, poly(ADP-ribose) polymerase, 1997), and here we provide an update that focuses on is a substrate for most caspases under somewhat exthe initiation, transmission, and regulation of caspase treme conditions in vitro, but in vivo is probably targeted activity. by caspases 3 and 7. Thus, by combining in vivo obserStructure, Mechanism, and Specificity vations with in vitro tests on suspect proteins, one can In common with other protease zymogens, generation recognize distinctions in substrate specificity within the of an active form requires limited proteolysis (Figure caspase family that fit remarkably closely to the S42S1 1). For the caspases, this results from cleavage in an subsite preferences deduced from synthetic peptidic interdomain linker segment to give a heterodimeric ensubstrates (Talanian et al., 1997; Thornberry et al., 1997) zyme, with both chains containing essential compo(Figure 2). nents of the catalytic machinery. Sometimes an N-termiSingularly important in this context is that caspase nal peptide, not required for enzyme activity, is released. zymogens are themselves substrates for caspases, and The role of the N peptide is only known for caspases 1 inspection of the individual interdomain linker in each and 8, where it behaves as a protein interaction domain zymogen reveals target sites that confirm the preference to modulate activation. of some caspases to activate others in a hierarchical The 3-D structures of caspases 1 and 3 reveal two relationship. Thus, pathways exist to transmit signals heterodimers interacting via the small chains, providing via sequential caspase activations, and this has been each molecule with two active sites. The primary recogmost extensively examined in apoptosis. nition pocket (S1) is well adapted to accept an Asp side Caspases Are Required for Apoptosis chain of the substrate, and additional pockets (S2–S4) A key to apoptosis is the discovery in many laboratories distinguish the caspases from each other (Figure 2). that, irrespective of the lethal stimulus, death results in Caspases cleave a number of cellular proteins, and