Dissecting the role of CENP-E in the maintenance of chromosome alignment

Abstract

The goal of mitosis is to ensure faithful segregation of genomic content into two identical daughter cells. As cells enter mitosis, the DNA becomes condensed into mitotic chromosomes, which must establish stable attachments to the mitotic spindle. Upon establishing proper kinetochore-microtubule attachments, chromosomes can then be guided to the center of the cell. Notably, this process of chromosome congression is a prerequisite for faithful chromosome segregation, since errors in this process lead to aneuploidy and chromosomal instability (CIN). One of the key players involved in chromosome congression is CENP-E, a kinesin-like motor protein that is enriched at kinetochores during mitosis. Despite mounting evidence demonstrating a role for CENP-E in chromosome congression, the importance of CENP-E activity once chromosomes are already aligned, is currently unknown. To this purpose, we first determined the optimal dose for inhibiting CENP-E function in both CIN and non-CIN human cell lines using three independent CENP-E inhibitors: PF-2771, GSK923295A and Compound A, which act through distinct allosteric mechanisms. Using high-throughput screening, we calculated the optimal doses required for partial and total CENP-E inhibition and then performed high-resolution live cell imaging to directly test CENP-E function in the maintenance of chromosome alignment. We show that CENP-E is essential for the maintenance of chromosome alignment in both CIN and non-CIN human cells. We find that relative sensitivity to CENP-E inhibitors is proportional for PF-2771 and GSK923295A when comparing CIN and non-CIN cell lines. However, diploid cells are less sensitive to CENP-E inhibition using Compound A, suggesting that this compound may be of higher therapeutic value for the treatment of cancer. Taken together, these results offer insight for our understanding of the therapeutic value of CENP-E inhibitors currently tested in clinical trials.