A novel role of farnesylation in targeting a mitotic checkpoint protein, human Spindly, to kinetochores.
Bottom Line: Inhibition of farnesylation using a farnesyl transferase inhibitor (FTI) abrogated hSpindly KT localization without affecting RZZ complex, CENP-E, and CENP-F KT localization.We showed that hSpindly is farnesylated in vivo and farnesylation is essential for its interaction with the RZZ complex and hence KT localization.Our data show a novel role of lipidation in targeting a checkpoint protein to KTs through protein-protein interaction.
Affiliation: Department of Oncology, University of Alberta, Edmonton, Alberta, Canada T6G 1Z2.Show MeSH
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Mentions: In our current structure function study, we mapped the KT localization domain of hSpindly to its C-terminal 294–605 aa (Fig. 8 A). Furthermore, our data showed that hSpindly undergoes farnesylation, which is essential for hSpindly KT localization. Substitution of the farnesylation motif with a geranylgeranylation motif does not support hSpindly KT localization. We found that FTI treatment does not interfere with KT localization of the RZZ complex, CENP-E, or CENP-F. Additionally, farnesylation plays a pivotal role in the interaction of hSpindly with the RZZ complex providing insight into lipid modification regulating checkpoint protein assembly. Our results showed that FTI treatment and hSpindly knockdown share the same phenotypes, prolonged prometaphase and metaphase with chromosome alignment defects, differing only in severity. Our analysis indicates that hSpindly is likely a key farnesylation target leading to FTI-induced mitotic defects.
Affiliation: Department of Oncology, University of Alberta, Edmonton, Alberta, Canada T6G 1Z2.