Network of protein interactions within the Drosophila inner kinetochore
Richter, Magdalena M
Royal Society Publishing
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Richter, M. M., Poznanski, J., Zdziarska, A., Czarnocki-Cieciura, M., Lipinszki, Z., Dadlez, M., Glover, D., & et al. (2016). Network of protein interactions within the Drosophila inner kinetochore. Open Biology, 6 (150238)https://doi.org/10.1098/rsob.150238
The kinetochore provides a physical connection between microtubules and the centromeric regions of chromosomes that is critical for their equitable segregation. The trimeric Mis12 sub-complex of the Drosophila kinetochore binds to the mitotic centromere using CENP-C as a platform. However, knowledge of the precise connections between Mis12 complex components and CENP-C has remained elusive despite the fundamental importance of this part of the cell division machinery. Here, we employ hydrogen–deuterium exchange coupled with mass spectrometry to reveal that Mis12 and Nnf1 form a dimer maintained by interacting coiled-coil (CC) domains within the carboxy-terminal parts of both proteins. Adjacent to these interacting CCs is a carboxy-terminal domain that also interacts with Nsl1. The amino-terminal parts of Mis12 and Nnf1 form a CENP-C-binding surface, which docks the complex and thus the entire kinetochore to mitotic centromeres. Mutational analysis confirms these precise interactions are critical for both structure and function of the complex. Thus, we conclude the organization of the Mis12–Nnf1 dimer confers upon the Mis12 complex a bipolar, elongated structure that is critical for kinetochore function.
chromosome, HDX-MS, kinetochore, centromere, mitosis, structure
The work was funded by the Foundation for Polish Science via an International PhD Projects Programme grant to MR and MD; Polish National Science Center via collaborative Harmonia 5 grant to MD and DMG (2013/10/M/NZ2/00298), and by the Medical Research Council and Cancer Research UK via programme grants to DMG. ZL was on leave from the Institute of Biochemistry, Biological Research Centre of the Hungarian Academy of Sciences, Szeged, Hungary, and supported by the FEBS Long Term fellowship. We are grateful to Magdalena Kaus-Drobek and Kinga Fituch for help with the structural characterisation of peptides. We thank Andrea Musacchio and his group for sharing their data before publication.
Cancer Research UK (18795)
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External DOI: https://doi.org/10.1098/rsob.150238
This record's URL: https://www.repository.cam.ac.uk/handle/1810/253630