Phosphorylation of targeting protein for Xenopus kinesin-like protein 2 (TPX2) at threonine 72 in spindle assembly.

The human ortholog of the targeting protein for Xenopus kinesin-like protein 2 (TPX2) is a cytoskeletal protein that plays a major role in spindle assembly and is required for mitosis. During spindle morphogenesis, TPX2 cooperates with Aurora A kinase and Eg5 kinesin to regulate microtubule organization. TPX2 displays over 40 putative phosphorylation sites identified from various high-throughput proteomic screenings. In this study, we characterize the phosphorylation of threonine 72 (Thr(72)) in human TPX2, a residue highly conserved across species. We find that Cdk1/2 phosphorylate TPX2 in vitro and in vivo. Using homemade antibodies specific for TPX2 phosphorylated at Thr(72), we show that this phosphorylation is cell cycle-dependent and peaks at M phase. Endogenous TPX2 phosphorylated at Thr(72) does not associate with the mitotic spindle. Furthermore, ectopic GFP-TPX2 T72A preferentially concentrates on the spindle, whereas GFP-TPX2 WT distributes to both spindle and cytosol. The T72A mutant also increases the proportion of cells with multipolar spindles phenotype. This effect is associated with increased Aurora A activity and abnormally elongated spindles, indicative of higher Eg5 activity. In summary, we propose that phosphorylation of Thr(72) regulates TPX2 localization and impacts spindle assembly via Aurora A and Eg5. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Citation

Su Yeon Shim, Ignacio Perez de Castro, Gernot Neumayer, Jian Wang, Sang Ki Park, Kamon Sanada, Minh Dang Nguyen. Phosphorylation of targeting protein for Xenopus kinesin-like protein 2 (TPX2) at threonine 72 in spindle assembly. The Journal of biological chemistry. 2015 Apr 03;290(14):9122-34

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PMID: 25688093

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