Featured Article - June 2012
Short description: The crystal structure of the mitotic checkpoint complex is reported.
Proper chromosome segregation is ensured in part by the mitotic checkpoint complex (MCC), which detects free chromosome ends and delays cell division until all chromosomes are attached to the mitotic spindle. This is called the spindle assembly checkpoint (SAC). The MCC implements the SAC by inhibiting the anaphase-promoting complex/cyclosome (APC/C), the E3 ubiquitin ligase involved in protein degradation required for chromosomes to segregate.
Barford and colleagues have taken a structural approach to studying the molecular machines involved in the SAC. They solved the crystal structure of the fission yeast MCC, consisting of the proteins Mad2, Mad3 and Cdc20, to 2.3Å resolution (PDB 4AEZ). The structure was obtained using the closed form of Mad2.
The MCC has a roughly triangular architecture. The helix–loop–helix motif of Mad3 interacts with Mad2 via its α-C helix and β8′–β8″ hairpin. This hairpin is known to undergo a substantial conformational change upon transition of Mad2 from the closed to the open state; the crystal structure thus satisfyingly reveals why it is only the closed form of Mad2 that can interact with Mad3.
Furthermore, the structure provides molecular insight into how Cdc20 recognizes APC/C substrates. Cdc20 is a co-activator of APC/C and recognizes its substrates via their KEN (Lys-Glu-Asn) box and D box motifs. In the context of the MCC, Mad2 and Mad3 function to inhibit Cdc20 substrate recognition. Mad3 has been proposed to be a pseudosubstrate inhibitor of Cdc20, blocking its access to APC/C substrates. Barford and colleagues thus studied the Mad3-Cdc20 interaction for clues into APC/C substrate recognition by co-activators.
They observed the KEN-box-binding site of Cdc20 on the top of its WD40 domain, and the D-box-binding site within a channel between blades 1 and 7 of the WD40 β-propeller. They confirmed the D-box-binding site by mutagenesis of the equivalent residues in the co-activator Cdh1 and used structural information about binding site disposition on Cdc20 to design peptide inhibitors of APC/C in complex with Cdh1. Finally, the authors docked the MCC structure into an electron microscopy-based molecular envelope model of APC/C-MCC to observe how Cdc20 substrate recognition is inhibited by Mad2 and Mad3 within the larger complex.
W.C.H. Chao et al. Structure of the mitotic checkpoint complex.
Nature 484, 208-l213 (2012). doi:10.1038/nature10896