WD40 domain of Apc1 is critical for the coactivator-induced allosteric transition that stimulates APC/C catalytic activity.
The anaphase-promoting complex/cyclosome (APC/C) is a large multimeric cullin-RING E3 ubiquitin ligase that orchestrates cell-cycle progression by targeting cell-cycle regulatory proteins for destruction via the ubiquitin proteasome system. The APC/C assembly comprises two scaffolding subcomplexes: the platform and the TPR lobe that together coordinate the juxtaposition of the catalytic and substrate-recognition modules. The platform comprises APC/C subunits Apc1, Apc4, Apc5, and Apc15. Although the role of Apc1 as an APC/C scaffolding subunit has been characterized, its specific functions in contributing toward APC/C catalytic activity are not fully understood. Here, we report the crystal structure of the N-terminal domain of human Apc1 (Apc1N) determined at 2.2-Å resolution and provide an atomic-resolution description of the architecture of its WD40 (WD40 repeat) domain (Apc1(WD40)). To understand how Apc1(WD40) contributes to APC/C activity, a mutant form of the APC/C with Apc1(WD40) deleted was generated and evaluated biochemically and structurally. We found that the deletion of Apc1(WD40) abolished the UbcH10-dependent ubiquitination of APC/C substrates without impairing the Ube2S-dependent ubiquitin chain elongation activity. A cryo-EM structure of an APC/C-Cdh1 complex with Apc1(WD40) deleted showed that the mutant APC/C is locked into an inactive conformation in which the UbcH10-binding site of the catalytic module is inaccessible. Additionally, an EM density for Apc15 is not visible. Our data show that Apc1(WD40) is required to mediate the coactivator-induced conformational change of the APC/C that is responsible for stimulating APC/C catalytic activity by promoting UbcH10 binding. In contrast, Ube2S activity toward APC/C substrates is not dependent on the initiation-competent conformation of the APC/C.
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom; Section of Structural Biology, Chester Beatty Laboratories, Institute of Cancer Research, London SW3 6JB, United Kingdom.
Qiuhong
Q
Li
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom;
Leifu
L
Chang
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom;
Shintaro
S
Aibara
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom;
Jing
J
Yang
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom;
Ziguo
Z
Zhang
Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom; dbarford@mrc-lmb.cam.ac.uk.
David
D
Barford
D000494:Allosteric Regulation / Q000235:genetics; D064173:Anaphase-Promoting Complex-Cyclosome / Q000737:chemistry* / Q000235:genetics; D015703:Antigens, CD; D064189:Apc1 Subunit, Anaphase-Promoting Complex-Cyclosome / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D015820:Cadherins / Q000737:chemistry* / Q000235:genetics; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics; D018360:Crystallography, X-Ray; D006801:Humans; D050505:Mutant Proteins / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D000072417:Protein Domains; D025801:Ubiquitin / Q000737:chemistry / Q000235:genetics; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000235:genetics; D044767:Ubiquitin-Protein Ligases / Q000235:genetics; D054875:Ubiquitination / Q000235:genetics; D000071678:WD40 Repeats / Q000235:genetics
C577335:ANAPC1 protein, human; C577350:ANAPC15 protein, human; D015703:Antigens, CD; D064189:Apc1 Subunit, Anaphase-Promoting Complex-Cyclosome; C513384:CDH1 protein, human; D015820:Cadherins; D018797:Cell Cycle Proteins; D050505:Mutant Proteins; D025801:Ubiquitin; C105156:UBE2C protein, human; C544962:Ube2S protein, human; D044763:Ubiquitin-Conjugating Enzymes; D064173:Anaphase-Promoting Complex-Cyclosome; C584471:CULL-RING ligase, human; D044767:Ubiquitin-Protein Ligases
APC/C; UbcH10; Ube2S; cell cycle; ubiquitination