Nonrandom γ-TuNA-dependent spatial pattern of microtubule nucleation at the Golgi
| Autor: | Roslin J. Thoppil, Irina Kaverina, Anna A. W. M. Sanders, Kevin Chang, Xiaodong Zhu, William R. Holmes |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Scaffold protein Nucleation Cell Culture Techniques Golgi Apparatus Cooperativity Spindle Apparatus Biology Microtubules 03 medical and health sciences symbols.namesake Microtubule Tubulin Humans Computer Simulation 14. Life underwater Molecular Biology Microtubule nucleation Centrosome Spatial Analysis food and beverages Microtubule organizing center Cell Biology Golgi apparatus 030104 developmental biology symbols Biophysics Brief Reports Microtubule-Associated Proteins Microtubule-Organizing Center |
| Zdroj: | Molecular Biology of the Cell |
| ISSN: | 1939-4586 |
| Popis: | GDMT (Golgi-derived microtubule) asymmetry is required for polarized cell motility, but its origin is elusive. Combining experimental and computational approaches, we find that GDMTs arise from spatially restricted hotspots that rely on γ-TuNA (γ-TuRC nucleation activator) activity. The nonrandom nucleation pattern underlies GDMT array asymmetry. Noncentrosomal microtubule (MT) nucleation at the Golgi generates MT network asymmetry in motile vertebrate cells. Investigating the Golgi-derived MT (GDMT) distribution, we find that MT asymmetry arises from nonrandom nucleation sites at the Golgi (hotspots). Using computational simulations, we propose two plausible mechanistic models of GDMT nucleation leading to this phenotype. In the “cooperativity” model, formation of a single GDMT promotes further nucleation at the same site. In the “heterogeneous Golgi” model, MT nucleation is dramatically up-regulated at discrete and sparse locations within the Golgi. While MT clustering in hotspots is equally well described by both models, simulating MT length distributions within the cooperativity model fits the data better. Investigating the molecular mechanism underlying hotspot formation, we have found that hotspots are significantly smaller than a Golgi subdomain positive for scaffolding protein AKAP450, which is thought to recruit GDMT nucleation factors. We have further probed potential roles of known GDMT-promoting molecules, including γ-TuRC-mediated nucleation activator (γ-TuNA) domain-containing proteins and MT stabilizer CLASPs. While both γ-TuNA inhibition and lack of CLASPs resulted in drastically decreased GDMT nucleation, computational modeling revealed that only γ-TuNA inhibition suppressed hotspot formation. We conclude that hotspots require γ-TuNA activity, which facilitates clustered GDMT nucleation at distinct Golgi sites. |
| Databáze: | OpenAIRE |
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