A novel mechanism that promotes mitotic spindle formation in cancer cells.


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Nikos Boutakoglou1, Eleni Petsalaki1, Sergio Lilla2, Sara Zanivan2, George Zachos1
1Other, 2Beatson Institute for Cancer Research

Abstract

Background

During cell division, the mitotic spindle consists mainly of microtubules (MTs) and is essential for accurate distribution of the genetic material to the two daughter cells. Errors in spindle formation can lead to incorrect separation of chromosomes that is associated with tumorigenesis or developmental disorders; however, the molecular mechanisms of mitotic spindle assembly are incompletely understood.


Method

To investigate Chk1 functions in the mitotic spindle, we used site directed mutagenesis, RNA silencing, replacement of endogenous proteins with transfected wild-type or mutant transgenes, in combination with confocal microscopy and biochemistry techniques.

Results

In the present study, we show that Chk1, a kinase involved in the cellular response to DNA damage, is essential for optimal density and effective polymerization of spindle MTs in human cells. Chk1 localizes to the centrosomes (the main centers of MT-organization in animal cells) in mitosis by confocal microscopy. Chk1 phosphorylates purified β-tubulin in kinase reactions in vitro at several conserved residues which were identified by mass spectrometry. Furthermore, reduced microtubule density in Chk1-deficient cells associates with formation of disorganized spindles. We propose that Chk1 phosphorylates β-tubulin to promote optimal spindle MT polymerization and spindle assembly.

Conclusion

In conclusion, these findings describe a novel mechanism that could protect against tumorigenesis, through regulating mitotic spindle formation.

Impact statement

These results describe novel mechanisms that could protect against aneuploidy and carcinogenesis by promoting proper spindle formation and accurate chromosome segregation during mitotic cell division.