Overexpression of EVI1 interferes with cytokinesis and leads to accumulation of cells with supernumerary centrosomes in G_0/1 phase

Abstract

Ectopic viral integration site 1 (EVI1), a transcription factor frequently overexpressed in myeloid neoplasias, has been implicated in the generation of malignancy-associated centrosomal aberrations and chromosomal instability. Here, we sought to investigate the underlying cause of centrosome amplification in EVI1-overexpressing cells. We found that overexpression of EVI1-HA in U2OS cells induced supernumerary centrosomes, which were consistently associated with enlarged nuclei or binuclear cells. Live cell imaging experiments identified cytokinesis failure as the underlying cause of this phenotype. In accordance with previous reports, EVI1 overexpression induced a partial cell cycle arrest in G_0/1 phase, accompanied by elevated cyclin D1 and p21 levels, reduced Cdk2 activity and activation of the p53 pathway. Supernumerary centrosomes predominantly occurred in resting cells, as identified by low levels of the proliferation marker K_i-67, leading to the conclusion that they result from tetraploidization after cytokinesis failure and are confined to G_0/1-arrested tetraploid cells. Depletion of p53 using siRNA revealed that further polyploidization of these cells was inhibited by the p53-dependent tetraploidy checkpoint.

Keywords: :

• EVI1
• chromosomal instability
• centrosome amplification
• mitosis
• cytokinesis

This article is referred to by:

New functions for ecotropic viral integration site 1 (EVI1), an oncogene causing aggressive malignant disease

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

Acknowledgments

We thank Manuel Grez for the EVI1 plasmid, Erich A. Nigg for the Cep170 antibody, Luigi Naldini for providing lentiviral packaging plasmids and the transfer vector pCCL.SIN.cPPT.SFFV.IRES.eGFP.wPRE and Sigrid Heil for excellent technical assistance. Support by the DKFZ Light Microscopy Facility and the Carl Zeiss Application Center, Heidelberg, is gratefully acknowledged. This work was supported by the Deutsche Forschungsgemeinschaft, grant SFB873, and by the Deutsche Krebshilfe, grants 108560, 10–1860-GI I and 107217.