MAD1: Kinetochore Receptors and Catalytic Mechanisms

Abstract

The mitotic checkpoint monitors kinetochore-microtubule attachment, delays anaphase onset and prevents aneuploidy when unattached or tensionless kinetochores are present in cells. Mitotic arrest deficiency 1 (MAD1) is one of the evolutionarily conserved core mitotic checkpoint proteins. MAD1 forms a cell cycle independent complex with MAD2 through its MAD2 interaction motif (MIM) in the middle region. Such a complex is enriched at unattached kinetochores and functions as an unusual catalyst to promote conformational change of additional MAD2 molecules, constituting a crucial signal amplifying mechanism for the mitotic checkpoint. Only MAD2 in its active conformation can be assembled with BUBR1 and CDC20 to form the Mitotic Checkpoint Complex (MCC), which is a potent inhibitor of anaphase onset. Recent research has shed light on how MAD1 is recruited to unattached kinetochores, and how it carries out its catalytic activity. Here we review these advances and discuss their implications for future research.

Keywords: MAD1; MAD2; kinetochore; mitosis; mitotic checkpoint; protein conformation.

Figure 1

Current model on MAD2 O-C conversion and MCC assembly. In prometaphase, unattached kinetochores initiate mitotic checkpoint signaling, leading to assembly of the mitotic checkpoint complex (MCC) consisting of BUBR1, CDC20, BUB3, and MAD2. The MCC inhibits APC/C ubiquitination activity until all chromosomes are correctly attached to microtubules. For MCC assembly, MAD2 needs to be converted from O conformation into C conformation.

Figure 2

Structural features of MAD1. (A) Non-coiled coil segments (ovals) are scatted along likely coiled-coil regions of MAD1 (in gray) as predicted by COILS program (Lupas et al., 1991). (B) Diagram of MAD1 showing its N-terminal domain (NTD), the domain containing the MAD2 interaction motif (MIM) and C-terminal domain (CTD). (C) Predicted model of MAD1^NTD dimer by Galaxy Homomer (Baek et al., 2017). (D) Crystal structure of MAD1^MIM (pink) dimer bound with two molecules of C-MAD2 (yellow) as in PDB 1GO4 (Sironi et al., 2002). Two O-MAD2 molecules (gray) from 2V64 (Mapelli et al., 2007) are also fitted. (E) Crystal structure of MAD1^CTD dimer from 4DZO (Kim et al., 2012). The RLK motifs are shown in red. Superscripts a&b denote two different chains of the same molecule. PyMol was used for structure visualization and model generation.

Figure 3

Phosphorylation (above) and interacting partners (below) of MAD1. Only the phosphorylation events and interactions with better-defined functional implications are shown. The interacting proteins are roughly grouped based on their binding to the NTD, MIM, or CTD of MAD1.

Figure 4

A proposed model on regulation of the catalytic activity of the MAD1:C-MAD2 heterotetramer to drive MAD2 O-C conversion. See text for details.