Phosphorylation of EB1 regulates the recruitment of CLIP-170 and p150glued to the plus ends of astral microtubules.

Jie Ran,Yijun Zhang,Min Liu,Miao Chen,Dengwen Li,Youguang Luo,Yang Yang,Jun Zhou

doi:10.18632/oncotarget.14222

Jie Ran, Yijun Zhang + Show 6 more

Open Access

https://doi.org/10.18632/oncotarget.14222

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Journal: Oncotarget	Publication Date: Dec 25, 2016
Citations: 14	License type: cc-by

Affiliation: Shandong Normal University, Nankai University

Abstract

Phosphorylation of end-binding protein 1 (EB1), a key member of microtubule plus end-tracking proteins (+TIPs), by apoptosis signal-regulating kinase 1 (ASK1) has been demonstrated to promote the stability of astral microtubules during mitosis by stimulating the binding of EB1 to microtubule plus ends. However, the roles of other members of the +TIPs family in ASK1/EB1-mediated regulation of astral microtubules are unknown. Herein, we show that ASK1-mediated phosphorylation of EB1 enhances the localization of cytoplasmic linker protein 170 (CLIP-170) and p150glued to the plus ends of astral microtubules. Depletion of ASK1 or expression of phospho-deficient or phospho-mimetic EB1 mutants results in changes in the levels of plus-end localized CLIP-170 or p150glued. Mechanistic studies reveal that EB1 phosphorylation promotes its interactions with CLIP-170 and p150glued, thereby recruiting these +TIPs to microtubules. Structural analysis suggests that serine-40 is the primary phosphorylation site on EB1 that exerts these effects. Together, these findings provide novel insight into the molecular mechanisms that regulate the interactions of EB1 with other +TIPs.

Highlights

During mitosis, accurate chromosome alignment requires dynamic attachment of sister kinetochores to spindle microtubules, in addition to the formation of interactions between astral microtubules and the cell cortex [1,2,3]
We have previously shown that overexpression of a triple phospho-mimetic end-binding protein 1 (EB1) mutant (S40D/T154D/ T206D; 3D), but not wild-type (WT) EB1 or a phosphodeficient mutant (S40A/T154A/T206A; 3A), rescued the loss of astral microtubules in cells depleted of apoptosis signal-regulating kinase 1 (ASK1) and EB1 [17]
These results suggest that ASK1-mediated phosphorylation of EB1 is required for astral microtubule stability

Summary

Introduction

Accurate chromosome alignment requires dynamic attachment of sister kinetochores to spindle microtubules, in addition to the formation of interactions between astral microtubules and the cell cortex [1,2,3]. To satisfy the mitotic spindle checkpoint and facilitate the onset of anaphase, the kinetochores and spindle microtubules and their associated proteins must be precisely integrated and organized, while astral microtubules need to be stabilized Defects in coordinating these events are associated with chromosome missegregation and genome aneuploidy, leading to cancer or developmental defects [1,2,3]. End-binding protein 1 (EB1), as a member of the microtubule plus end-tracking proteins (+TIPs), plays a central role in the regulation of microtubule dynamics by recruiting other +TIPs and binding partners to microtubule plus ends [4,5,6] One such +TIP is cytoplasmic linker protein 170 (CLIP-170), which associates with EB1 and tubulin to dynamically track plus ends of growing microtubules and positively regulate microtubule growth [4, 7]. CLIP-170 can form a complex with other proteins that links microtubules to the cell cortex in order to respond to cell polarity cues [11]

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Conclusion