Welburn lab paper featured in Journal of Cell Science. Authors Image Legal and co-workers defined the molecular requirement for the BubR1-dependent recruitment of CENP-E to kinetochores. They show a acidic stretch of amino acids in CENP-E is essential for the interaction with the pseudokinase domain of BubR1. Their work highlighted that while another unknown pathway support additional recruitment of CENP-E, BubR1 is essential for the CENP-E dependent chromosome alignment and biorientation. Legal, T., Hayward, D., Gluszek-Kustusz, A., Blackburn, E.A., Spanos, C., Rappsilber, J., Gruneberg, U., and Welburn, J.P.I. Summary of Paper by Lori Koch DNA is packaged into chromosomes which must be replicated and segregated into new cells during cell division. Each chromosome associates with a protein complex called the kinetochore to connect to the dynamic microtubule fibres of the mitotic spindle that drive segregation. The molecular motor protein CENP-E associates with unattached kinetochores on chromosomes to facilitate their proper alignment on spindle microtubules. In their recent work published in the Journal of Cell Science, PhD student Thibault Legal and their colleagues in Julie Welburn’s group determined the molecular requirements for the localization of CENP-E at kinetochores in human cells. By performing truncation analyses, they determined a short region of the CENP-E C-terminus that was sufficient for localization to kinetochores. They generated recombinant CENP-E2055-2068 and performed pull-down experiments with cell lysate to identify which proteins interacted with it. This identified the spindle checkpoint protein BubR1 as a major binding partner. Using size-exclusion chromatography (SEC), they showed that the CENP-E C-terminus interacts with the C-terminus of BubR1, especially when CENP-E forms a dimer. Interestingly, isothermal calorimetry (ITC) showed that a dimeric CENP-E construct and the BubR1 pseudo-kinase domain associated at a stoichiometry of 1:1, suggesting that one dimeric CENP-E motor can associate with two molecules of BubR1. Finally, they determined that depletion of BubR1 in cells caused a decrease of CENP-E from kinetochores during checkpoint activation. Together, their results suggest that CENP-E binds to unattached kinetochores to facilitate their alignment through interaction with the spindle checkpoint protein BubR1. Related links Journal Link Welburn Lab Website DOI This article was published on 2024-06-17
