State of the APC/C: organization, function, and structure

Abstract

The ubiquitin-proteasome protein degradation system is involved in many essential cellular processes including cell cycle regulation, cell differentiation, and the unfolded protein response. The anaphase-promoting complex/cyclosome (APC/C), an evolutionarily conserved E3 ubiquitin ligase, was discovered 15 years ago because of its pivotal role in cyclin degradation and mitotic progression. Since then, we have learned that the APC/C is a very large, complex E3 ligase composed of 13 subunits, yielding a molecular machine of approximately 1 MDa. The intricate regulation of the APC/C is mediated by the Cdc20 family of activators, pseudosubstrate inhibitors, protein kinases and phosphatases and the spindle assembly checkpoint. The large size, complexity, and dynamic nature of the APC/C represent significant obstacles toward high-resolution structural techniques; however, over the last decade, there have been a number of lower resolution APC/C structures determined using single particle electron microscopy. These structures, when combined with data generated from numerous genetic and biochemical studies, have begun to shed light on how APC/C activity is regulated. Here, we discuss the most recent developments in the APC/C field concerning structure, substrate recognition, and catalysis.

Figure 1

Theoretical model of APC/C ubiquitination pathway. The four APC/C modules (Catalytic=Apc2, Apc11; Scaffold = Apc1, Apc4, Apc5; TPR=Apc3, Apc6, Apc8, Cdc26, Apc10, and species-specific subunits; and Activator=Cdc20 family member) are labeled. El = Ub-activating enzyme, E2 = Ub-conjugating enzyme, IR = IR motif in Cdc20 family members, C-box= conserved motif in Cdc20 family members, degrons=APC/C degradation motifs (e.g., D box, KEN box), DUB = deubiquitinating enzyme, * indicates elongation E2.

Figure 2

Structural analysis of the APC/C spanning from yeasts to vertebrates. (A) ~20Å structure of the Saccharomyces cerevisiae APC/C purified from asynchronous cells (Passmore et al., 2005) (EMDB1174). (B) Single-particle cryo-EM analysis of the mitotic Schizosaccharomyces pombe APC/C^Cdc20 (Ohi et al., 2007). Position of APC/C subunits and the activator Cdc20, found by antibody labeling and difference mapping, respectively, are labeled. Cut4 corresponds to human Apc1, Nuc2 corresponds to human Apc3, and Lid1 corresponds to human Apc4. (C) ~24Å structure of the Xenopus APC/C. Approximate locations of recombinant Cdh1 are labeled (Herzog et al., 2009). (D) Human APC/C (~26Å) purified from asynchronous cells (EMDB 1139) (Dube et al., 2005). (E) Human apo-APC/C (~20 Å) purified from checkpoint active lysates (EMDB 1592) (Herzog et al., 2009). Position of APC/C subunits as well as the activators Cdc20 and Cdh1, localized using antibody labeling and recombinant proteins, respectively, are noted. (F) ~20Å structure of the human APC/C bound to the MCC inhibitory complex (APC/C^MCC, EMDB 1591) (Herzog et al., 2009). Position of MCC labeled in dark gray. Images for (D) and (F) were kindly provided by Franz Herzog (IMP-Vienna), Jan-Michael Peters (IMP-Vienna), and Holger Stark (Max Planck Institute). Scale bar for all panels, 5nm.