PXD065516

PXD065516 is an original dataset announced via ProteomeXchange.

Title	Cryo-EM structures of the Plant Augmin reveal its intertwined coiled-coil assembly, antiparallel dimerization and NEDD1 binding mechanisms
Description	Microtubule (MT) branch nucleation is fundamental for building parallel MT networks in eukaryotic cells. In plants and metazoans, MT branch nucleation requires Augmin and NEDD1 proteins which bind along MTs and then recruit and activate the gamma-tubulin ring complex (g-TuRC). Augmin is a fork-shaped assembly composed of eight coiled-coil subunits, while NEDD1 is a b-propellor protein that bridges across MTs, Augmin, and g-TuRC during MT branch nucleation. Here, we reconstitute hetero-tetrameric and hetero-octameric Arabidopsis thaliana Augmin assemblies, resolve their subunit interactions using crosslinking mass spectrometry and determine 3.7 to 7.3-Å cryo-EM structures for the V-junction and extended regions of Augmin. These structures allowed us to generate a complete de novo plant Augmin model that reveals the long-range multi coiled-coil interfaces that stabilize its 40-nm hetero-octameric fork-shaped organization. We discovered the dual calponin homology (CH) domain forming its MT binding site at one end of its V-junction can assume open and closed conformations. We determined a 12-Å dimeric Augmin cryo-EM structure revealing Augmin undergoes anti-parallel dimerization through two conserved surfaces along Augmin’s extended region. We reconstituted the NEDD1 b-propellor with Augmin revealing it directly binds into its V-junction and enhances Augmin dimerization. Our studies suggest that cooperativity between the Augmin dual CH domains and NEDD1 binding site may regulate Augmin V-junction dual binding to MT lattices. This unique V-shaped dual binding and organization anchors Augmins along MTs generating a platform to recruit g-TuRC and activate branched MT nucleation.
HostingRepository	PRIDE
AnnounceDate	2025-10-31
AnnouncementXML	Submission_2025-10-31_13:50:15.621.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD065516
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Yuqi Tang
SpeciesList	scientific name: Arabidopsis thaliana (Mouse-ear cress); NCBI TaxID: NEWT:3702;
ModificationList	monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Q Exactive HF; Orbitrap Ascend

Revision	Datetime	Status	ChangeLog Entry
0	2025-06-26 07:24:16	ID requested
⏵ 1	2025-10-31 13:50:16	announced

10.6019/PXD065516;

submitter keyword: microtubule, augmin, g-TuRC, XL-MS, NEDD1

Stephen D. Fried
contact affiliation	Department of Chemistry, Department of Biology (by courtesy), Thomas C.Jenkins Department of Biophysics (by courtesy), Johns Hopkins University
contact email	sdfried@jhu.edu
lab head
Yuqi Tang
contact affiliation	Johns Hopkins University
contact email	ytang93@jh.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD065516
     1. Label: PRIDE project
     2. Name: Cryo-EM structures of the Plant Augmin reveal its intertwined coiled-coil assembly, antiparallel dimerization and NEDD1 binding mechanisms