Abstract
Author SummaryMicrotubules are polymers of tubulin that undergo successive cycles of growth and shrinkage so that the cell can maintain a stable yet adaptable cytoskeleton. In neurons, the microtubule motor protein dynein and its partner complex dynactin drive retrograde transport along microtubules from the distal axon towards the cell body. In addition to binding to dynein, the p150Glued subunit of dynactin independently binds directly to microtubules. We hypothesized that by binding to microtubules, p150Glued might also alter microtubule dynamics. We demonstrate using biochemistry and microscopy in vitro and in cells that p150Glued stabilizes microtubules by inhibiting the transition from growth to shrinkage. We show that specific domains of p150Glued encoded by neuronally enriched splice-forms are necessary for this activity. Although depletion of p150Glued in nonpolarized cells does not alter microtubule dynamics, depletion of endogenous p150Glued in neurons leads to dramatic microtubule instability. Strikingly, a mutation in p150Glued known to cause the neurodegenerative disorder Perry syndrome abolishes this activity. In summary, we identified a previously unappreciated function of dynactin in direct regulation of the microtubule cytoskeleton. This activity may enhance generic microtubule stability in the cell, but could be especially important in specific areas of the cell where dynactin and dynein are loaded onto microtubules.
Highlights
Microtubules are dynamic, polarized polymers of tubulin that serve as tracks for long-distance transport in eukaryotic cells
Depletion of p150Glued in nonpolarized cells does not alter microtubule dynamics, depletion of endogenous p150Glued in neurons leads to dramatic microtubule instability
Hydrodynamic analysis indicated that both polypeptides are highly elongated (Rs/Rmin.1.9, Figure S1B), consistent with previous electron microscopy images of dynactin [22] that indicate that the p150Glued dimer projects outward from the Arp1 filament that forms the base of the dynactin complex
Summary
Microtubules are dynamic, polarized polymers of tubulin that serve as tracks for long-distance transport in eukaryotic cells. Transport along microtubules is especially important yet vulnerable to disruption, as these cells are long-lived and postmitotic with elongated axonal processes that can extend up to a meter [1]. In large, postmitotic cells like neurons, noncentrosomal nucleation may be important [5,6,7]. The dynamics of polymerization and depolymerization are strongly influenced by microtubule-associated proteins (MAPs). A spatially specialized group of MAPs that localize to the microtubule plus end, the plus end-tracking proteins (+TIPs), are ideally poised to modulate dynamics in cells [8]
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