TTBK2 with EB1/3 regulates microtubule dynamics in migrating cells through KIF2A phosphorylation.
Bottom Line: TTBK2 depletion reduced MT lifetime (facilitated shrinkage and suppressed rescue) and impaired HeLa cell migration, and these phenotypes were partially restored by KIF2A co-depletion.Expression of nonphosphorylatable KIF2A, but not wild-type KIF2A, reduced MT lifetime and slowed down the cell migration.These findings indicate that TTBK2 with EB1/3 phosphorylates KIF2A and antagonizes KIF2A-induced depolymerization at MT plus ends for cell migration.
Affiliation: Department of Cell Pharmacology, Nagoya University Graduate School of Medicine, Showa, Nagoya 466-8550, Japan.Show MeSH
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Mentions: Finally, we examined whether TTBK2 regulates cell migration via KIF2A because appropriate MT dynamics and integrity are required for cell migration (Watanabe et al., 2005; Kuijpers and Hoogenraad, 2011; Etienne-Manneville, 2013). We performed wound healing assays using HeLa cells. The control HeLa cells migrated toward the gap during wound healing, but TTBK2 depletion inhibited this cell migration (Fig. 7 A). Tracking of each cell demonstrated the directional and persistent migration of control cells, whereas TTBK2-depleted cells migrated less efficiently, including a loss of directionality toward the wound (Fig. 7 B). When quantifying migratory properties (Fig. 7 C), control HeLa cells moved at a velocity of 14.1 ± 3.25 µm/h for a total distance of 125 ± 35.8 µm from their original position (n > 150). TTBK2 depletion decreased the velocity to 10.4 ± 3.48 µm/h and the total distance to 77.4 ± 37.4 µm (siTTBK2-1; n > 150). Essentially, similar results were obtained using a different TTBK2 siRNA (unpublished data). The expression of resTTBK2-WT rescued the inhibitory effects of TTBK2 depletion on HeLa cell migration (Fig. 7 C and Fig. S4 A), establishing the essential role of TTBK2 in these migratory properties. We then examined the effects of KIF2A depletion on cell migration and found that KIF2A depletion also inhibited the migration (Fig. 7, B and C; and Fig. S4 A). TTBK2 and KIF2A double depletion, however, partially rescued the migration defects caused by TTBK2 or KIF2A single depletion (Fig. 7, B and C; and Fig. S4 A). Considering that TTBK2 inactivates KIF2A, these results indicate that the proper activity level of KIF2A is required for cell migration, and activities of TTBK2 and KIF2A should be tightly balanced for effective cell migration.
Affiliation: Department of Cell Pharmacology, Nagoya University Graduate School of Medicine, Showa, Nagoya 466-8550, Japan.