A quantitative systems view of the spindle assembly checkpoint

Abstract

The idle assembly checkpoint acts to delay chromosome segregation until all duplicated sister chromatids are captured by the mitotic spindle. This pathway ensures that each daughter cell receives a complete copy of the genome. The high fidelity and robustness of this process have made it a subject of intense study in both the experimental and computational realms. A significant number of checkpoint proteins have been identified but how they orchestrate the communication between local spindle attachment and global cytoplasmic signalling to delay segregation is not yet understood. Here, we propose a systems view of the spindle assembly checkpoint to focus attention on the key regulators of the dynamics of this pathway. These regulators in turn have been the subject of detailed cellular measurements and computational modelling to connect molecular function to the dynamics of spindle assembly checkpoint signalling. A review of these efforts reveals the insights provided by such approaches and underscores the need for further interdisciplinary studies to reveal in full the quantitative underpinnings of this cellular control pathway.

Figure 1

Schematic view of spindle assembly checkpoint signalling. (A) Cells enter mitosis with unattached kinetochores that actively generate inhibitory signals (strong red alarm signal) to prevent APC/C activation. This stabilizes the high levels of cyclin B and securin that prevent anaphase onset. (B) Attachment of spindle microtubules to unattached kinetochores locally turns off kinetochore-mediated inhibition, but cytoplasmic inhibition, potentially diminished, is still supported by other unattached kinetochores (weaker red signal). The progressive attachment of microtubules generates a weak signal in the cytoplasm that promotes the disengagement of the checkpoint (weak green alarm signal) (C) Capture of all chromosomes results in the complete loss of signal generation from kinetochores (weakest red signal), permitting the greater relief of inhibition on the APC/C in the cytoplasm (stronger green alarm). Activation of the APC/C promotes the destabilization of cyclin B and securin. (D) Sufficient loss of substrates (cyclin B and securin) promotes the activation of separase and cleavage of the cohesins permitting the onset of anaphase and segregation of the sister chromatids.

Figure 2

Modular organization of the spindle assembly checkpoint. (A) The interactions between the modules that comprise the spindle assembly checkpoint. The K-microtubule module represents the proteins at the kinetochore that control microtubule attachment. The K-checkpoint module, the network of proteins at the kinetochores that generate the inhibitory flux of Mad2:Cdc20 and A^*. Finally, the cytoplasm module represents the reactions of MCC:APC/C association and release taking place in the cytoplasm—the balance between APC/C inhibition and its release. The filled arrows represent the molecular interactions controlling the activity of the scaffold at the unattached kinetochore. The open arrows indicate net fluxes, which result in the generation of Mad2:Cdc20 complexes from the unattached kinetochores and the release of free Cdc20 through active dissociation within the cytoplasm. (B) The spindle assembly checkpoint signalling elements of the kinetochore (K-Checkpoint) catalyse, through the Mad1:Mad2 scaffold, the formation of Mad2:Cdc20 complexes. In this representation, the kinetochore can also modulate the level of cytoplasmic MCC:APC/C dissociation activity through the proposed A to A^* modification. Red complexes act to inhibit APC/C activity, whereas green activate. (C) The microtubule attachment module of the kinetochore (K-Microtubule) involves the microtubule attachment complex, Ndc80, and the Dynein-binding protein Spindly. The action of Spindly and the Rod–Zw10–Zwilch complex (RZZ) controls indirectly, through microtubule attachment, kinetochore-mediated inhibitor generation. (D) The cytoplasm has three submodules. The first forms the MCC:APC/C inhibitory complex from the Mad2:Cdc20 complex and other cytoplasmic components (BubR1:Bub3 and APC/C). The second actively dissociates the inhibited APC/C into the component parts, through the activity of A. Note that A^* (inactive) is converted to A (active) in the cytoplasmic module. The third component comprises non-kinetochore mechanisms for generation of the Mad2:Cdc20 complex, specifically through cytoplasmic amplification from Closed Mad2 complexes in the cytoplasm (Mad2:Cdc20, MCC (not shown) and MCC:APC/C). This last reaction is represented with a dashed line.

Figure 3

Molecular interpretation of wiring diagrams proposed in biophysical models. (A) In the simplest model, ‘direct inhibition model' in Doncic et al (2005), the checkpoint (here Cdc20—active) is inhibited (Cdc20i—inactive) by direct contact with the kinetochore (black dot in the figure). In red, inhibited species, in green, the active ones. (B) According to the ‘indirect inhibition model' (Doncic et al, 2005), kinetochores produces an active species (Mad2^*) that inhibits Cdc20 through a complex that is dissociated in the cytoplasm. (C) The ‘Mad2 template model' postulates that the role of the kinetochore in (B) can be played by the complex Mad2:Cdc20 in the cytoplasm. In this description, we include the activated species Mad2^*, which is missing in the original formulation of the template model (De Antoni et al, 2005). The combination of (B, C) gives rise to the full template model. Of note is the autocatalytic loop that is present in (C).