title: A proteomic study of mitotic phase-specific interactors of EB1 reveals a role for SXIP-mediated protein interactions in anaphase onset.
abstract: Microtubules execute diverse mitotic events that are spatially and temporally separated; the underlying regulation is poorly understood. By combining drug treatments, large-scale immunoprecipitation and mass spectrometry, we report the first comprehensive map of mitotic phase-specific protein interactions of the microtubule-end binding protein, EB1. EB1 interacts with some, but not all, of its partners throughout mitosis. We show that the interaction of EB1 with Astrin-SKAP complex, a key regulator of chromosome segregation, is enhanced during prometaphase, compared to anaphase. We find that EB1 and EB3, another EB family member, can interact directly with SKAP, in an SXIP-motif dependent manner. Using an SXIP defective mutant that cannot interact with EB, we uncover two distinct pools of SKAP at spindle microtubules and kinetochores. We demonstrate the importance of SKAP's SXIP-motif in controlling microtubule growth rates and anaphase onset, without grossly disrupting spindle function. Thus, we provide the first comprehensive map of temporal changes in EB1 interactors during mitosis and highlight the importance of EB protein interactions in ensuring normal mitosis.
authors.affiliation: Department of Genetics, University of Cambridge, Cambridge, UK.
authors.forename: Naoka
authors.initials: N
authors.lastname: Tamura
authors.affiliation: Department of Genetics, University of Cambridge, Cambridge, UK Present address: European Research Institute for the Biology of Ageing, University of Groningen, Groningen, Netherlands.
authors.forename: Judith E
authors.initials: JE
authors.lastname: Simon
authors.affiliation: Department of Genetics, University of Cambridge, Cambridge, UK Present address: Institute for Biochemistry II, Goethe University Frankfurt am Main, Germany.
authors.forename: Arnab
authors.initials: A
authors.lastname: Nayak
authors.affiliation: Department of Genetics, University of Cambridge, Cambridge, UK.
authors.forename: Rajesh T
authors.initials: RT
authors.lastname: Shenoy
authors.affiliation: Keio University, Yokohama, Japan.
authors.forename: Noriko
authors.initials: N
authors.lastname: Hiroi
authors.affiliation: Department of Genetics, University of Cambridge, Cambridge, UK.
authors.forename: Viviane
authors.initials: V
authors.lastname: Boilot
authors.affiliation: Keio University, Yokohama, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Funahashi
authors.affiliation: Department of Genetics, University of Cambridge, Cambridge, UK vmd20@cam.ac.uk.
authors.forename: Viji M
authors.initials: VM
authors.lastname: Draviam
keywords: Cell cortex; Kinetochore; Microtubule; Mitosis; Plus-end