NuMA regulates mitotic spindle assembly, structural dynamics and function via phase separation

Mengjie Sun,Wangfei Chi,Biying Yang,Chuanmao Zhang,Qing Jiang,He Ren,Guangwei Xin,Mingkang Jia

doi:10.1038/s41467-021-27528-6

Abstract

A functional mitotic spindle is essential for accurate chromosome congression and segregation during cell proliferation; however, the underlying mechanisms of its assembly remain unclear. Here we show that NuMA regulates this assembly process via phase separation regulated by Aurora A. NuMA undergoes liquid-liquid phase separation during mitotic entry and KifC1 facilitates NuMA condensates concentrating on spindle poles. Phase separation of NuMA is mediated by its C-terminus, whereas its dynein-dynactin binding motif also facilitates this process. Phase-separated NuMA droplets concentrate tubulins, bind microtubules, and enrich crucial regulators, including Kif2A, at the spindle poles, which then depolymerizes spindle microtubules and promotes poleward spindle microtubule flux for spindle assembly and structural dynamics. In this work, we show that NuMA orchestrates mitotic spindle assembly, structural dynamics and function via liquid-liquid phase separation regulated by Aurora A phosphorylation.

Highlights

A functional mitotic spindle is essential for accurate chromosome congression and segregation during cell proliferation; the underlying mechanisms of its assembly remain unclear
We reveal that the binding of NuMA droplets to microtubules facilitates sorting of acentrosomal microtubule asters into the spindle microtubule array for the mitotic spindle assembly and structural dynamics
To investigate the molecular mechanisms underlying mitotic spindle assembly, structural dynamics, and function, we first depleted NuMA in HCT-116 cells by using the auxininducible degron (AID) system, which knocks in a specific vector, mAID-mClover-FLAG, that fuses to endogenous NuMA (NuMA-mACF) to promote efficient endogenous NuMA protein degradation under doxycycline (Dox) and 3-indoleacetic acid (IAA) induction[16] and investigated the consequences of NuMA depletion

Summary

Introduction

A functional mitotic spindle is essential for accurate chromosome congression and segregation during cell proliferation; the underlying mechanisms of its assembly remain unclear. Phase-separated NuMA droplets concentrate tubulins, bind microtubules, and enrich crucial regulators, including Kif2A, at the spindle poles, which depolymerizes spindle microtubules and promotes poleward spindle microtubule flux for spindle assembly and structural dynamics. We reveal that the binding of NuMA droplets to microtubules facilitates sorting of acentrosomal microtubule asters into the spindle microtubule array for the mitotic spindle assembly and structural dynamics These data provide a molecular mechanism for spatiotemporal regulation of spindle assembly, structural dynamics, and function mediated by NuMA phase separation and this work may have more implications for understanding the mechanism of accurate chromosome distribution into two daughter cells regulated by NuMA during the cell division

Methods

Results

Conclusion