Oncogene (2010) 29, 4752–4765 & 2010 Macmillan Publishers Limited All rights reserved 0950-9232/10 www.nature.com/onc REVIEW New insights into apoptosis signaling by Apo2L/TRAIL F Gonzalvez and A Ashkenazi Department of Molecular Oncology, Genentech Inc., South San Francisco, CA, USA Apoptosis ligand 2 tumor necrosis factor (TNF)-related important component in neurodegenerative disorders apoptosis-inducing ligand (Apo2L/TRAIL) belongs to a such as Alzheimer’s and Huntington’s disease (Rohn small subset of proapoptotic protein ligands in the TNF et al., 2001; Sanchez Mejia and Friedlander, 2001), superfamily. This subset, which also includes Fas ligand stroke (Prunell et al., 2005), ischemic heart disease and TNF-a, can activate the extrinsic apoptotic cell death (Hayakawa et al., 2002), AIDS (Casella and Finkel, pathway on binding to cognate death receptors at the cell 1997) and infertility (Said et al., 2004). In mammalian surface. Over the past 10 years, Apo2L/TRAIL has cells, apoptosis induction occurs through two distinct emerged as a promising candidate for cancer therapy, on pathways: intrinsic and extrinsic. The intrinsic pathway the basis of its unique ability to trigger apoptosis in is activated by intracellular events and depends on the various types of cancer cells without significant toxicity release of proapoptotic factors from the mitochondria toward normal cells. Herein, we review key advances in (Danial and Korsmeyer, 2004). The extrinsic pathway understanding the molecular events that control apoptosis receives signals through the binding of extracellular signaling by Apo2L/TRAIL, which may aid in the protein ligands to proapoptotic death receptors (DRs), development of cancer therapies based on the extrinsic located on the cell surface (Ashkenazi and Dixit, 1998). apoptotic pathway. Both pathways lead to hierarchical activation of Oncogene (2010) 29, 4752–4765; doi:10.1038/onc.2010.221; specialized cysteine–aspartate proteases called caspases published online 7 June 2010 (Lavrik et al., 2005b; Li and Yuan, 2008). Apoptotic signals first activate initiator caspases, including cas- Keywords: DR4; DR5; glycosylation; caspase-8; cullin pase-2, -8, -9 and -10, by inducing their oligomerization 3; p62 at multiprotein platforms (Chen et al., 2002; Boatright et al., 2003; Bouchier-Hayes et al., 2009). Once stimulated, initiator caspases proteolytically activate Introduction the downstream effector caspases, including caspase-3, -6 and -7, which in turn cleave numerous essential Apoptosis is a major mechanism of programmed cell cellular proteins, thereby leading to the unique death used by metazoan organisms to eliminate super- morphological and biochemical features of apoptosis, fluous and irreparably damaged cells (Danial and such as plasma membrane ‘blebbing’, cell shrinkage, Korsmeyer, 2004). It has a crucial role in shaping chromatin condensation and internucleosomal DNA organs during development and controls homeostasis fragmentation. and integrity of tissues throughout life (Meier et al., The intrinsic pathway involves mitochondrial outer- 2000; Chowdhury et al., 2006). Apoptosis can be membrane permeabilization (MOMP) and consequent triggered by a wide variety of stimuli, including release of apoptogenic factors, such as cytochrome c, developmental cues, severe cellular stress or damage to AIF and Smac-DIABLO, from the mitochondria into essential cellular components, caused by heat shock, the cytoplasm (Green and Reed, 1998; Kroemer et al., radiation, cytotoxic drugs, infection and oncogenic 1998; Wang, 2001). Cell-intrinsic stimuli control MOMP transformation. Aberrant regulation of apoptotic cell through the proteins of the Bcl-2 family (van Loo et al., death mechanisms is an important pathological factor in 2002). Members of this family are evolutionarily a variety of major human diseases (Fadeel et al., 1999). conserved and can be divided into three subgroups, Deficiency in apoptosis is one of the key hallmarks of according to function or structural similarity within four cancer (Hanahan and Weinberg, 2000) and also Bcl-2-homology domains (designated BH1–4) (Petros contributes to certain autoimmune diseases (Prasad et al., 2004; Youle and Strasser, 2008). The BH3-only and Prabhakar, 2003) and metabolic disorders (McKen- proapoptotic proteins are activated by transcriptional or zie et al., 2004). In contrast, excessive apoptosis is an post-translational mechanisms in response to specific cellular stress conditions (for example, Bik by endo- plasmic reticular stress, Bim and Bad by cytokine Correspondence: Dr A Ashkenazi, Department of Molecular Oncol- deprivation, Puma and Noxa by DNA damage). Once ogy, Genentech Inc., 1 DNA Way, South San Francisco, CA 94080, activated, BH3-only proteins promote oligomerization USA. E-mail: [email protected] of the multi-BH domain proapoptotic proteins Bax and Received 22 February 2010; revised 30 April 2010; accepted 4 May 2010; Bak in the mitochondrial outer membrane, thereby published online 7 June 2010 inducing MOMP. In contrast, the multi-BH domain Apoptosis signaling by Apo2L/TRAIL F Gonzalvez and A Ashkenazi 4753 antiapoptotic members, such as Bcl-2, Bcl-XL and Mcl-1, DRs inhibit MOMP induction by neutralizing specific proapoptotic family members. The release of apopto- DRs: distinct members of the TNFR superfamily genic factors from the mitochondria causes numerous DRs belong to the TNF receptor (TNFR) superfamily, cellular changes that commit the cell to death and is which comprises more than 20 proteins, involved in therefore considered ‘the point of no return’ in the diverse biological functions ranging from the regulation intrinsic pathway (Green and Amarante-Mendes, 1998). of cell death or survival to differentiation and In the cytosol, cytochrome c induces the assembly of a immunity (Nagata, 1997; Ashkenazi and Dixit, 1998; multimeric complex called the apoptosome, containing Krammer, 2000). Similar to other TNFRs, DRs are type the adaptor protein Apaf-1 and the apoptosis-initiating I transmembrane proteins that contain cysteine-rich protease caspase-9. The apoptosome mediates the extracellular domains (CRDs) (Figure 1). They are clustering and activation of caspase-9 (Li et al., 1997), uniquely characterized, however, by a relatively con- which goes on to process and activates effector caspases served cytoplasmic ‘death domain’ (DD) of B80 amino (Rodriguez and Lazebnik, 1999). The X chromosome- acids, which is essential for transmitting the apoptotic linked inhibitor of apoptosis protein (XIAP) provides an signal. To date, six DRs have been identified: important checkpoint to prevent inadvertent caspase TNFR1 (TNFRSF1A), Fas (also known as CD95, activation: XIAP binds to caspases-3, -7 and -9 through APO-1 or TNFRSF6), DR3 (TNFRSF12), DR4 (also its baculovirus IAP repeat domains and inhibits caspase known as TRAILR1 or TNFRSF10A), DR5 (also activity (Deveraux et al., 1998; Salvesen and Duckett, known as TRAILR2 or TNFRSF10B) and DR6 2002; Eckelman et al., 2006; Varfolomeev and Vucic, (TNFRSF21) (Figure 1). Extensive studies on TNFR1, 2008). However, release of mitochondrial Smac, which Fas, DR4 and DR5, with their respective cognate antagonizes XIAP, can override this checkpoint (Verha- ligands, TNF-a (TNFSF2), Fas ligand (FasL or gen et al., 2000; Srinivasula et al., 2001). TNFSF6) and Apo2L/TRAIL (TNFSF10), have been The extrinsic pathway transmits signals from extra- reported in the literature. DR6 has recently emerged cellular ligands through proapoptotic DRs to the apopto- as a crucial component of a neuronal self-destruction tic caspase machinery (Ashkenazi and Dixit, 1998). pathway associated with Alzheimer’s disease Depending on cell type, the extrinsic and intrinsic path- (Nikolaev et al., 2009). The extracellular fragment of ways may converge (Ozoren and El-Deiry, 2002; Barnhart the b-amyloid precursor protein (N-APP), characteristic et al., 2003; LeBlanc and Ashkenazi, 2003). In mammals, to Alzheimer’s brain pathology, binds DR6 and the extrinsic pathway has an important role in regulating triggers neuronal degeneration through activation of various aspects of immunity. Under physiological condi- caspase-6. tions, it participates in selection and maintenance of the On ligation, DRs recruit DD-binding adaptor pro- immune repertoire and regulates the immune response teins—either Fas-associated DD (FADD) or TNFR- (Opferman and Korsmeyer, 2003; Wilson et al., 2009). associated DD (TRADD)—which then promote the Furthermore, the extrinsic pathway helps remove infected, assembly of multiprotein signaling complexes. Although transformed or damaged cells. Natural killer cells and all DRs possess a DD, they can be distinguished by cytotoxic T lymphocytes express proapoptotic ligands of the pivotal adaptor protein they recruit and the the TNF superfamily, which recognize specific DRs on the consequent signaling complex they assemble. Fas, surface of target cells. Binding of these ligands to their DR4 and DR5 typically bind to FADD to form the cognate DRs results in the assembly of a death-inducing DISC and activate apoptosis (Algeciras-Schimnich signaling complex (DISC). The DISC stimulates the et al., 2003; Lavrik et al., 2005a; Wilson et al., 2009). initiator proteases caspase-8 and -10, thereby driving In contrast, TNFR1 and DR3 usually recruit TRADD downstream effector caspase activation and apoptosis to assemble distinct signaling complexes that regulate (Peter and Krammer, 2003). cellular responses such as survival, differentiation or Among the proapoptotic