Abstract
ABSTRACTDynein is the sole processive minus-end-directed microtubule motor found in animals. It has roles in cell division, membrane trafficking, and cell migration. Together with dynactin, dynein regulates centrosomal orientation to establish and maintain cell polarity, controls focal adhesion turnover and anchors microtubules at the leading edge. In higher eukaryotes, dynein/dynactin requires additional components such as Bicaudal D to form an active motor complex and for regulating its cellular localization. Spindly is a protein that targets dynein/dynactin to kinetochores in mitosis and can activate its motility in vitro. However, no role for Spindly in interphase dynein/dynactin function has been found. We show that Spindly binds to the cell cortex and microtubule tips and colocalizes with dynein/dynactin at the leading edge of migrating U2OS cells and primary fibroblasts. U2OS cells that lack Spindly migrated slower in 2D than control cells, although centrosome polarization appeared to happen properly in the absence of Spindly. Re-expression of Spindly rescues migration, but the expression of a mutant, which is defective for dynactin binding, failed to rescue this defect. Taken together, these data demonstrate that Spindly plays an important role in mediating a subset of dynein/dynactin's function in cell migration.
Highlights
Cell migration is required for development and homeostasis in almost all multi-cellular organisms
When we used an affinity-purified antibody raised against the full-length recombinant protein to stain U2OS cells that were grown in a monolayer and scratched to induce cell migration, we noticed that, in addition to the expected nuclear staining, there was a cytoplasmic pool of protein (Fig. 1A, upper)
To examine Spindly’s localization in a more migratory cell type and to determine if it localizes with any components of the dynein/ dynactin complex, we fixed and stained primary human fibroblasts
Summary
Cell migration is required for development and homeostasis in almost all multi-cellular organisms. Cells lacking Spindly are slow to migrate in a scratch-wound assay, a defect that can be rescued by the reintroduction of the wild-type protein but not by the expression of a mutant that fails to bind to dynactin.
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