Cdk1-mediated DIAPH1 phosphorylation maintains metaphase cortical tension and inactivates the spindle assembly checkpoint at anaphase

Koutarou Nishimura,Shige H Yoshimura,Midori Shimada,Narumi Suzuki,Makoto Nakanishi,Keiko Kono,Yoshie Chiba,Zixian Jiang,Toru Hirota,Yoshikazu Johmura,Katashi Deguchi,Kazuhiko S K Uchida

doi:10.1038/s41467-019-08957-w

Abstract

Animal cells undergo rapid rounding during mitosis, ensuring proper chromosome segregation, during which an outward rounding force abruptly increases upon prometaphase entry and is maintained at a constant level during metaphase. Initial cortical tension is generated by the actomyosin system to which both myosin motors and actin network architecture contribute. However, how cortical tension is maintained and its physiological significance remain unknown. We demonstrate here that Cdk1-mediated phosphorylation of DIAPH1 stably maintains cortical tension after rounding and inactivates the spindle assembly checkpoint (SAC). Cdk1 phosphorylates DIAPH1, preventing profilin1 binding to maintain cortical tension. Mutation of DIAPH1 phosphorylation sites promotes cortical F-actin accumulation, increases cortical tension, and delays anaphase onset due to SAC activation. Measurement of the intra-kinetochore length suggests that Cdk1-mediated cortex relaxation is indispensable for kinetochore stretching. We thus uncovered a previously unknown mechanism by which Cdk1 coordinates cortical tension maintenance and SAC inactivation at anaphase onset.

Highlights

Animal cells undergo rapid rounding during mitosis, ensuring proper chromosome segregation, during which an outward rounding force abruptly increases upon prometaphase entry and is maintained at a constant level during metaphase
The formin homology 1 (FH1) domain is required for the interaction with the actin monomer-binding protein profilin, whereas the formin homology 2 (FH2) domain is responsible for actin filament nucleation[17]
A clear mobility shift of 3×FLAG-DIAPH1 bands was detected in HeLa cells synchronized with nocodazole and was reversed with calf intestine alkaline phosphatase (CIP) (Fig. 1b), indicating that the mobility shift of DIAPH1 was due to its phosphorylation

Summary

Introduction

Animal cells undergo rapid rounding during mitosis, ensuring proper chromosome segregation, during which an outward rounding force abruptly increases upon prometaphase entry and is maintained at a constant level during metaphase. We demonstrate here that Cdk1-mediated phosphorylation of DIAPH1 stably maintains cortical tension after rounding and inactivates the spindle assembly checkpoint (SAC). Thereafter, the cortex tension is maintained at a constant level during metaphase under the progressive accumulation of myosin II but with a decrease in actin thickness[14] This is somewhat surprising since RhoA is activated at the cortex during early mitosis[23], raising the expectation that DIAPH1-dependent F-actin would progressively accumulate on the cortex and the tension would increase. We found that Cdk[1] phosphorylated DIAPH1, which inhibited the interaction between DIAPH1 and profilin[1] (PFN1) during metaphase This inhibition is required for maintaining the cortical tension at a constant level and for the proper inactivation of the SAC at the onset of anaphase

Methods

Results

Conclusion