Abstract
Proper segregation of chromosomes depends on a functional spindle assembly checkpoint (SAC) and requires kinetochore localization of the Bub1 and Mad1/Mad2 checkpoint proteins. Several aspects of Mad1/Mad2 kinetochore recruitment in human cells are unclear and in particular the underlying direct interactions. Here we show that conserved domain 1 (CD1) in human Bub1 binds directly to Mad1 and a phosphorylation site exists in CD1 that stimulates Mad1 binding and SAC signalling. Importantly, fusion of minimal kinetochore-targeting Bub1 fragments to Mad1 bypasses the need for CD1, revealing that the main function of Bub1 is to position Mad1 close to KNL1 MELT repeats. Furthermore, we identify residues in Mad1 that are critical for Mad1 functionality, but not Bub1 binding, arguing for a direct role of Mad1 in the checkpoint. This work dissects functionally relevant molecular interactions required for spindle assembly checkpoint signalling at kinetochores in human cells.
Highlights
Proper segregation of chromosomes depends on a functional spindle assembly checkpoint (SAC) and requires kinetochore localization of the Bub[1] and Mad1/mitotic arrest deficient (Mad)[2] checkpoint proteins
We show here that conserved domain 1 (CD1) is required for a phosphoregulated direct interaction between human Bub[1] and Mad[1] and that disturbance of this interaction is detrimental to the SAC
The function of CD1 is unclear, while the ABBA motif is required for recruiting a fraction of Cdc[20] to kinetochores (we will discuss recent parallel data on CD1 from the Yu lab published during the reviewing of this manuscript in the discussion; Supplementary Fig. 1 and refs 31,32)
Summary
Proper segregation of chromosomes depends on a functional spindle assembly checkpoint (SAC) and requires kinetochore localization of the Bub[1] and Mad1/Mad[2] checkpoint proteins. Accurate segregation of sister chromatids during cell division depends on the spindle assembly checkpoint (SAC), which in response to improper attachments between kinetochores and microtubules generates a diffusible ‘wait anaphase’ inhibitor[1,2,3] This inhibitor is the mitotic checkpoint complex (MCC) composed of the Mad[2] and BubR1–Bub[3] checkpoint proteins bound to Cdc[20], the mitotic co-activator of the anaphase-promoting complex (APC/C)[4]. The exact mechanism behind recruitment of the Mad1/Mad[2] complex to kinetochores in humans has not been established, but given the central role of this complex for SAC signalling this is crucial to understand This contrasts the situation in budding yeast and worms where a direct interaction between Mad[1] and Bub[1] has been shown to localize Mad1/Mad[2] to kinetochores[37,38,39]. The exact contribution of Bub[1] and the RZZ complex is still to be fully dissected
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
