Cdk1-phosphorylated CUEDC2 promotes spindle checkpoint inactivation and chromosomal instability

Abstract

Aneuploidy and chromosomal instability are major characteristics of human cancer. These abnormalities can result from defects in the spindle assembly checkpoint (SAC), which is a surveillance mechanism for accurate chromosome segregation through restraint of the activity of the anaphase-promoting complex/cyclosome (APC/C). Here, we show that a CUE-domain-containing protein, CUEDC2, is a cell-cycle regulator that promotes spindle checkpoint inactivation and releases APC/C from checkpoint inhibition. CUEDC2 is phosphorylated by Cdk1 during mitosis. Depletion of CUEDC2 causes a checkpoint-dependent delay of the metaphase–anaphase transition. Phosphorylated CUEDC2 binds to Cdc20, an activator of APC/C, and promotes the release of Mad2 from APC/C–Cdc20 and subsequent APC/C activation. CUEDC2 overexpression causes earlier activation of APC/C, leading to chromosome missegregation and aneuploidy. Interestingly, CUEDC2 is highly expressed in many types of tumours. These results suggest that CUEDC2 is a key regulator of mitosis progression, and that CUEDC2 dysregulation might contribute to tumour development by causing chromosomal instability.

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Figure 1: The Cdk1-dependent phosphorylation of CUEDC2 is required for the metaphase–anaphase transition.
Figure 2: The knockdown of CUEDC2 did not affect mitotic spindle, BubR1 and Mad2 signals at kinotechores.
Figure 3: CUEDC2 regulates APC/C-mediated ubiquitylation and degradation depending on SAC.
Figure 4: The interaction of CUEDC2 with Cdc20 depends on the phosphorylation at Ser 110, but not the CUE domain.
Figure 5: CUEDC2 is required for the disassociation of Mad2 from APC/C–Cdc20 complex.
Figure 6: Overexpression of CUEDC2 leads to chromosome missegregation and aneuploidy.

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Acknowledgements

We thank H. T. Yu and Y. X. Zheng for discussions, M. W. Kirschner, T. Wu and G. W. Fang for technical help, Z. G. Liu for reading, S. Doxsey for HeLa cells stably expressing GFP–H2B and hTERT-RPE1 cells, E. D. Salmon for GFP–Cdc20 expression plasmid, Y. Liu for pFlag-Cdk1-AF (constitutively active) and pFlag-Cdk1-DN (kinase dead) plasmids, J. Pines for pVenus-N1 Cyclin B1 plasmid, M. W. Kirschner for the plasmids coding securin and geminin, P. K. Jackson for the plasmid pCS2-Cyclin B1, R. Benezra for the plasmid pFlag-CMV2-Mad2, H. T. Yu for pCS2-securin and pCS2-securin (ΔDB) plasmids and H. M. Wang for capturing tissue array images. This work was supported by the National Natural Science Foundation of China (no 91029733; no 30830097; no 30871234; no 30872348; no 81025010; no 30900754), the National High Technology Research and Development Program of China (2009AA02Z103 and 2009ZX09503-001) and the National Basic Research Program of China (2010CB911900 and 2010CB529904).

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X-M.Z. and H-Y.L. supervised the project; Y-F.G., Teng L. and Yan C. designed and carried out most of the experiments; Y-F.G. and Y-B.W. contributed to chromosome spread analysis; W-H.L. and K.H. analysed the phosphorylation modification with mass spectrometry; W-N.Z. and R.M. carried out immunohistochemistry analysis; Y-B.W., Tao L. and C.Z. contributed to the preparation of complementary DNA vector constructs; M.Y., Yuan C. and R.M. prepared the CUEDC2 antibody; W-L.G., B.L. and L.C. developed stable cell lines; J-H.M., Q.X. and X.P. carried out the statistics; T.Z., A-L.L., X-M.Z. and H-Y.L. analysed the data; Y-F.G., Teng L., Yan C., H-Y.L. and X-M.Z. wrote the paper.

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Correspondence to Hui-Yan Li or Xue-Min Zhang.

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Gao, YF., Li, T., Chang, Y. et al. Cdk1-phosphorylated CUEDC2 promotes spindle checkpoint inactivation and chromosomal instability. Nat Cell Biol 13, 924–933 (2011). https://doi.org/10.1038/ncb2287

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