Regulators of IAP function: coming to grips with the grim reaper Andreas Bergmann  , Amy Yi-Pei Yang and Mayank Srivastava Inhibitor of apoptosis proteins (IAPs) are a conserved class of proteins that control apoptosis in both vertebrates and invertebrates. They exert their anti-apoptotic function through inhibition of caspases, the principal executioners of apoptotic cell death. Recent advances in vertebrates and Drosophila have demonstrated that IAPs use ubiquitin conjugation to control the stability, and thus the activity, of select target proteins. The Drosophila IAP1 gene is an instructive example: it employs at least two distinct ubiquitin-dependent mechanisms of protein destruction. The apoptosis-inducing genes grim, reaper and hid modulate these mechanisms, and determine the outcome. Addresses The University of Texas MD Anderson Cancer Center, Department of Biochemistry & Molecular Biology, Unit 117, 1515 Holcombe Blvd, Houston, TX 77030 USA  e-mail: andreas@bergman.net Current Opinion in Cell Biology 2003, 15:717–724 This review comes from a themed issue on Cell division, growth and death Edited by Jonathon Pines and Sally Kornbluth 0955-0674/$ – see front matter ß 2003 Elsevier Ltd. All rights reserved. DOI 10.1016/j.ceb.2003.10.002 Abbreviations BIR baculovirus IAP repeat DIAP1 Drosophila inhibitor of apoptosis protein 1 GMR glass-multimer reporter IAP inhibitor of apoptosis protein RHG reaper, hid and grim RING really interesting new gene UBC ubiquitin conjugating XIAP X-linked IAP Introduction Apoptosis is a physiological cell-suicide process that plays an important role during the development of multicellular organisms and is critical for the maintenance of tissue homeostasis. Caspases, a highly specialized class of cell- death proteases, are the main executioners of apoptosis [1,2]. They are synthesized as inactive zymogen precur- sors and require proteolytic cleavage for activation. In this process, the prodomain is cleaved off, and a large and small subunit are generated [1,2]. Activation of upstream caspases such as Caspase-9 is regulated by cytochrome c and Apaf-1. Upstream caspases activate downstream cas- pases in an amplifying cascade, cleaving one another in sequence. These modes of caspase activation have been extensively reviewed [1,2]. However, recent advances in Drosophila have indicated that the inhibition of caspases is a highly dynamic process involving protein–protein inter- actions and proteolytic degradation. The Drosophila inhi- bitor of apoptosis protein 1 (Diap1) plays a central role in this regulation, and is the focus of this review. The apoptotic machinery is conserved between verte- brates and Drosophila, and there are fly homologs of caspases, Bcl-2 family members, Apaf-1 and IAPs [3]. Genetic analysis in Drosophila has identified three addi- tional genes that are essential for embryonic cell death in this species. These genes are reaper, hid and grim, and are collectively referred to as the RHG genes [4–6]. Two more genes, sickle and jafrac2, with similar characteristics to the reaper, hid and grim genes, have recently been identified [7–10]. Chromosomal deletions removing the RHG genes completely block apoptosis during embry- ogenesis, and cause embryonic lethality [4], demonstrat- ing the importance of the RHG genes for apoptosis in Drosophila. The RHG genes encode novel proteins without signif- icant homology to other proteins in the database. How- ever, they share a common motif at the N terminus (Figure 1b,c). This motif, referred to as the RHG motif [11], is essential for the ability of the RHG proteins to induce apoptosis: truncation of the motif results in partial or complete loss of the apoptosis-inducing activity of the RHG proteins. Recently, two mammalian factors, Smac/ Diablo and Omi/HtrA2, have been identified that behave functionally as homologs of the RHG genes and possess the RHG motif (Figure 1c) [12–14]. Interestingly, the RHG motif has to be present at the extreme N terminus of the proteins. Drosophila Reaper, Hid, Grim and Sickle carry the RHG motif immediately following the initiator methionine [4–9] (Figure 1c). However, the RHG motif of Smac/Diablo, Omi/HtrA2 and Drosophila Jafrac2 is not located at the N terminus of the precursor forms of these proteins, and requires proteolytic processing for N- terminal exposure. Strikingly, because they are mitochon- drial proteins, but encoded in the nucleus, the RHG motif is revealed at the N terminus after their mitochon- drial import when the signal sequence is cleaved off [10,12–14]. A third mammalian protein containing a RHG motif is Caspase-9 (Figure 1c). Again, after proteo- lytic processing of Caspase-9, the small subunit exposes a tetrapeptide motif at the N terminus that functions as a RHG [15]. Overexpression of the Drosophila RHG genes in flies or in mammalian cell culture is sufficient to induce apoptosis [5,6,16,17]. For example, transgenes that place the 717 www.current-opinion.com Current Opinion in Cell Biology 2003, 15:717–724