Structures of APC/CCdh1 with substrates identify Cdh1 and Apc10 as the D-box co-receptor

Abstract

The ubiquitylation of cell cycle regulatory proteins by the large multimeric anaphase promoting complex (APC/C) controls sister chromatid segregation and the exit from mitosis 1,2. Selection of APC/C targets is achieved through recognition of destruction motifs, predominantly the D-box 3 and KEN-box 4. Although this process is known to involve a co-activator protein (either Cdc20 or Cdh1) together with core APC/C subunits 1,2, the structural basis for substrate recognition and ubiquitylation is not understood. Here, we investigated the APC/C using single particle electron microscopy (EM) and determined a cryo-EM map of APC/CCdh1 bound to a D-box peptide at ~10 Å resolution. We find that a combined catalytic and substrate recognition module is located within the central cavity of the APC/C assembled from Cdh1, Apc10 - a core APC/C subunit previously implicated in substrate recognition 5,6,7, and the cullin domain of Apc2. Cdh1 and Apc10, identified from difference maps, create a co-receptor for D-box following repositioning of Cdh1 towards Apc10. Using NMR spectroscopy we demonstrate specific D-box – Apc10 interactions, consistent with a role for Apc10 in directly contributing towards D-box recognition by the APC/CCdh1 complex. Our results rationalise the contribution of both co-activator and core APC/C subunits to D-box recognition 8,9 and provide a structural framework for understanding mechanisms of substrate recognition and catalysis by the APC/C.