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- Publisher Website: 10.1083/jcb.202009146
- Scopus: eid_2-s2.0-85100459712
- PMID: 33496729
- WOS: WOS:000626353200018
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Article: Biochemical reconstitutions reveal principles of human γ-TuRC assembly and function
Title | Biochemical reconstitutions reveal principles of human γ-TuRC assembly and function |
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Authors | |
Keywords | Biochemistry Cytoskeleton Structural Biology |
Issue Date | 2021 |
Publisher | Rockefeller University Press. The Journal's web site is located at https://jcb.rupress.org/ |
Citation | Journal of Cell Biology, 2021, v. 220 n. 3, p. article no. e202009146 How to Cite? |
Abstract | The formation of cellular microtubule networks is regulated by the γ-tubulin ring complex (γ-TuRC). This ∼2.3 MD assembly of >31 proteins includes γ-tubulin and GCP2-6, as well as MZT1 and an actin-like protein in a “lumenal bridge” (LB). The challenge of reconstituting the γ-TuRC has limited dissections of its assembly and function. Here, we report a biochemical reconstitution of the human γ-TuRC (γ-TuRC-GFP) as a ∼35 S complex that nucleates microtubules in vitro. In addition, we generate a subcomplex, γ-TuRCΔLB-GFP, which lacks MZT1 and actin. We show that γ-TuRCΔLB-GFP nucleates microtubules in a guanine nucleotide–dependent manner and with similar efficiency as the holocomplex. Electron microscopy reveals that γ-TuRC-GFP resembles the native γ-TuRC architecture, while γ-TuRCΔLB-GFP adopts a partial cone shape presenting only 8–10 γ-tubulin subunits and lacks a well-ordered lumenal bridge. Our results show that the γ-TuRC can be reconstituted using a limited set of proteins and suggest that the LB facilitates the self-assembly of regulatory interfaces around a microtubule-nucleating “core” in the holocomplex. |
Persistent Identifier | http://hdl.handle.net/10722/305738 |
ISSN | 2021 Impact Factor: 8.077 2020 SCImago Journal Rankings: 5.414 |
PubMed Central ID | |
ISI Accession Number ID |
DC Field | Value | Language |
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dc.contributor.author | Wieczorek, M | - |
dc.contributor.author | Ti, SC | - |
dc.contributor.author | Urnavicius, L | - |
dc.contributor.author | Molloy, KR | - |
dc.contributor.author | Aher, A | - |
dc.contributor.author | Chait, BT | - |
dc.contributor.author | Kapoor, TM | - |
dc.date.accessioned | 2021-10-20T10:13:38Z | - |
dc.date.available | 2021-10-20T10:13:38Z | - |
dc.date.issued | 2021 | - |
dc.identifier.citation | Journal of Cell Biology, 2021, v. 220 n. 3, p. article no. e202009146 | - |
dc.identifier.issn | 0021-9525 | - |
dc.identifier.uri | http://hdl.handle.net/10722/305738 | - |
dc.description.abstract | The formation of cellular microtubule networks is regulated by the γ-tubulin ring complex (γ-TuRC). This ∼2.3 MD assembly of >31 proteins includes γ-tubulin and GCP2-6, as well as MZT1 and an actin-like protein in a “lumenal bridge” (LB). The challenge of reconstituting the γ-TuRC has limited dissections of its assembly and function. Here, we report a biochemical reconstitution of the human γ-TuRC (γ-TuRC-GFP) as a ∼35 S complex that nucleates microtubules in vitro. In addition, we generate a subcomplex, γ-TuRCΔLB-GFP, which lacks MZT1 and actin. We show that γ-TuRCΔLB-GFP nucleates microtubules in a guanine nucleotide–dependent manner and with similar efficiency as the holocomplex. Electron microscopy reveals that γ-TuRC-GFP resembles the native γ-TuRC architecture, while γ-TuRCΔLB-GFP adopts a partial cone shape presenting only 8–10 γ-tubulin subunits and lacks a well-ordered lumenal bridge. Our results show that the γ-TuRC can be reconstituted using a limited set of proteins and suggest that the LB facilitates the self-assembly of regulatory interfaces around a microtubule-nucleating “core” in the holocomplex. | - |
dc.language | eng | - |
dc.publisher | Rockefeller University Press. The Journal's web site is located at https://jcb.rupress.org/ | - |
dc.relation.ispartof | Journal of Cell Biology | - |
dc.rights | Journal of Cell Biology. Copyright © Rockefeller University Press. | - |
dc.rights | ©2021 AUTHOR et al. Originally published in JOURNAL NAME. https://doi.org/10.1083/jcb.202009146 | - |
dc.subject | Biochemistry | - |
dc.subject | Cytoskeleton | - |
dc.subject | Structural Biology | - |
dc.title | Biochemical reconstitutions reveal principles of human γ-TuRC assembly and function | - |
dc.type | Article | - |
dc.identifier.email | Ti, SC: jeffti@hku.hk | - |
dc.identifier.authority | Ti, SC=rp02617 | - |
dc.description.nature | published_or_final_version | - |
dc.identifier.doi | 10.1083/jcb.202009146 | - |
dc.identifier.pmid | 33496729 | - |
dc.identifier.pmcid | PMC7844428 | - |
dc.identifier.scopus | eid_2-s2.0-85100459712 | - |
dc.identifier.hkuros | 327595 | - |
dc.identifier.volume | 220 | - |
dc.identifier.issue | 3 | - |
dc.identifier.spage | article no. e202009146 | - |
dc.identifier.epage | article no. e202009146 | - |
dc.identifier.isi | WOS:000626353200018 | - |
dc.publisher.place | United States | - |