Three BUB1 and BUBR1/ MAD3-related spindle assembly checkpoint proteins are required for accurate mitosis in Arabidopsis.

The spindle assembly checkpoint ( SAC) is a refined surveillance mechanism which ensures that chromosomes undergoing mitosis do not segregate until they are properly attached to the spindle microtubules ( MT). The SAC has been extensively studied in metazoans and yeast, but little is known about its role in plants., We identified proteins interacting with a MT-associated protein MAP65-3, which plays a critical role in organising mitotic MT arrays, and carried out a functional analysis of previously and newly identified SAC components., We show that Arabidopsis SAC proteins BUB3.1, MAD2, BUBR1/ MAD3s and BRK1 interact with each other and with MAP65-3. We found that two BUBR1/ MAD3s interacted specifically at centromeres. When stably expressed in Arabidopsis, BRK1 localised to the kinetochores during all stages of the mitotic cell cycle. Early in mitosis, BUB3.1 and BUBR1/ MAD3.1 localise to the mitotic spindle, where MAP65-3 organises spindle MTs. A double-knockout mad3.1 mad3.2 mutant presented spindle MT abnormalities, chromosome misalignments on the metaphase plate and the production of lagging chromosomes and micronuclei during mitosis., We conclude that BRK1 and BUBR1/ MAD3-related proteins play a key role in ensuring faithful chromosome segregation during mitosis and that their interaction with MAP65-3 may be important for the regulation of MT-chromosome attachment. [ABSTRACT FROM AUTHOR]