Cyclin B1 scaffolds MAD1 at the kinetochore corona to activate the mitotic checkpoint.

Lindsey A Allan,Adrian T Saurin,Andrea Musacchio,Magda Camacho Reis,Geert Jpl Kops,Pim J Huis In ‘T Veld,Sabine Wohlgemuth,Giuseppe Ciossani

doi:10.15252/embj.2019103180

Lindsey A Allan, Adrian T Saurin + Show 6 more

Open Access

https://doi.org/10.15252/embj.2019103180

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Abstract

Cyclin B:CDK1 is the master kinase regulator of mitosis. We show here that, in addition to its kinase functions, mammalian Cyclin B also scaffolds a localised signalling pathway to help preserve genome stability. Cyclin B1 localises to an expanded region of the outer kinetochore, known as the corona, where it scaffolds the spindle assembly checkpoint (SAC) machinery by binding directly to MAD1. In vitro reconstitutions map the key binding interface to a few acidic residues in the N‐terminal region of MAD1, and point mutations in this sequence abolish MAD1 corona localisation and weaken the SAC. Therefore, Cyclin B1 is the long‐sought‐after scaffold that links MAD1 to the corona, and this specific pool of MAD1 is needed to generate a robust SAC response. Robustness arises because Cyclin B1:MAD1 localisation loses dependence on MPS1 kinase after the corona has been established, ensuring that corona‐localised MAD1 can still be phosphorylated when MPS1 activity is low. Therefore, this study explains how corona‐MAD1 generates a robust SAC signal, and it reveals a scaffolding role for the key mitotic kinase, Cyclin B1:CDK1, which ultimately helps to inhibit its own degradation.

Highlights

During mitosis, all duplicated chromosomes must attach correctly to microtubules so they can segregate properly when the cell divides
Immunofluorescence analysis demonstrated that this localisation pattern reflects specific binding to unattached kinetochores, which is reminiscent of the checkpoint protein MAD1 (Fig 1B and C)
Cyclin B1 depends on MPS1 activity to be established at this location, but thereafter it became largely insensitive to MPS1 inhibition (Fig 1D and E), as shown previously for MAD1 (Hewitt et al, 2010; Etemad et al, 2019)

Summary

Introduction

All duplicated chromosomes must attach correctly to microtubules so they can segregate properly when the cell divides. This attachment is mediated via the kinetochore, which is a giant molecular complex assembled on chromosomes at the centromere (Musacchio & Desai, 2017). As well as attaching to microtubules, the kinetochore must regulate this process to ensure it occurs correctly. One aspect of this regulation involves the activation of the mitotic checkpoint, otherwise known as the spindle assembly checkpoint (SAC), which blocks mitotic exit until all kinetochores have attached to microtubules. The generation of MCC is so efficient that every single kinetochore signalling centre must eventually be extinguished by microtubule attachment to allow the cell to exit mitosis (Rieder et al, 1995; Dick & Gerlich, 2013)

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