Emi1 stably binds and inhibits the anaphase-promoting complex/cyclosome as a pseudosubstrate inhibitor.
The periodic destruction of mitotic cyclins is triggered by the activation of the anaphase-promoting complex/cyclosome (APC/C) in mitosis. Although the ability of the APC/C to recognize destruction box (D-box) substrates oscillates throughout the cell cycle, the mechanism regulating APC/C binding to D-box substrates remains unclear. Here, we show that the APC/C inhibitor Emi1 tightly binds both the APC/C and its Cdh1 activator, binds to the D-box receptor site on the APC/C(Cdh1), and competes with APC/C substrates for D-box binding. Emi1 itself contains a conserved C-terminal D-box, which provides APC/C-binding affinity, and a conserved zinc-binding region (ZBR), which antagonizes APC/C E3 ligase activity independent of tight APC binding. Mutation of the ZBR converts Emi1 into a D-box-dependent APC/C substrate. The identification of a direct Emi1-APC/C complex further explains how Emi1 functions as a stabilizing factor for cyclin accumulation and the need to destroy Emi1 for APC/C activation in mitosis. The combination of a degron/E3 recognition site and an anti-ligase function in Emi1 suggests a general model for how E3 substrates evolve to become pseudosubstrate inhibitors.
Miller
Julie J
JJ
Program in Molecular Pharmacology, Stanford University School of Medicine, Stanford, California 94305, USA.
Summers
Matthew K
MK
Hansen
David V
DV
Nachury
Maxence V
MV
Lehman
Norman L
NL
Loktev
Alex
A
Jackson
Peter K
PK
D020816:Amino Acid Motifs; D064173:Anaphase-Promoting Complex-Cyclosome; D015703:Antigens, CD; D001667:Binding, Competitive; D015820:Cadherins / Q000378:metabolism / Q000502:physiology; D018797:Cell Cycle Proteins / Q000378:metabolism / Q000502:physiology*; D002467:Cell Nucleus / Q000201:enzymology / Q000378:metabolism; D017124:Conserved Sequence; D004791:Enzyme Inhibitors / Q000378:metabolism; D044783:F-Box Proteins / Q000378:metabolism / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D007399:Interphase / Q000502:physiology; D011485:Protein Binding; D013379:Substrate Specificity; D043743:Ubiquitin-Protein Ligase Complexes / Q000037:antagonists & inhibitors* / Q000378:metabolism*
D015703:Antigens, CD; C513384:CDH1 protein, human; D015820:Cadherins; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D044783:F-Box Proteins; C498960:FBXO5 protein, human; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome