Cargando…
Reconstitution and mechanistic dissection of the human microtubule branching machinery
Branching microtubule (MT) nucleation is mediated by the augmin complex and γ-tubulin ring complex (γ-TuRC). However, how these two complexes work together to promote this process remains elusive. Here, using purified components from native and recombinant sources, we demonstrate that human augmin a...
Autores principales: | , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Rockefeller University Press
2022
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9129923/ https://www.ncbi.nlm.nih.gov/pubmed/35604367 http://dx.doi.org/10.1083/jcb.202109053 |
Sumario: | Branching microtubule (MT) nucleation is mediated by the augmin complex and γ-tubulin ring complex (γ-TuRC). However, how these two complexes work together to promote this process remains elusive. Here, using purified components from native and recombinant sources, we demonstrate that human augmin and γ-TuRC are sufficient to reconstitute the minimal MT branching machinery, in which NEDD1 bridges between augmin holo complex and GCP3/MZT1 subcomplex of γ-TuRC. The single-molecule experiment suggests that oligomerization of augmin may activate the branching machinery. We provide direct biochemical evidence that CDK1- and PLK1-dependent phosphorylation are crucial for NEDD1 binding to augmin, for their synergistic MT-binding activities, and hence for branching MT nucleation. In addition, we unveil that NEDD1 possesses an unanticipated intrinsic affinity for MTs via its WD40 domain, which also plays a pivotal role in the branching process. In summary, our study provides a comprehensive understanding of the underlying mechanisms of branching MT nucleation in human cells. |
---|