PXD036295

PXD036295 is an original dataset announced via ProteomeXchange.

Title	Analyzing the mechanisms that facilitate the subtype-specific assembly of γ-aminobutyric acid type A receptors
Description	Impaired inhibitory signaling underlies the pathophysiology of many neuropsychiatric and neurodevelopmental disorders including autism spectrum disorders and epilepsy. Neuronal inhibition is regulated by synaptic and extrasynaptic γ-aminobutyric acid type A receptors (GABAARs), which mediate phasic and tonic inhibition, respectively. These two GABAAR subtypes differ in their function, ligand sensitivity, and physiological properties. Importantly, they contain different α subunit isoforms: synaptic GABAARs contain the α1-3 subunits whereas extrasynaptic GABAARs contain the α4-6 subunits. While the subunit composition is critical for the distinct roles of synaptic and extrasynaptic GABAAR subtypes in inhibition, the molecular mechanism of the subtype-specific assembly has not been elucidated. To address this issue, we purified endogenous α1- and α4-containing GABAARs from adult murine forebrains and examined their subunit composition and interacting proteins using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and quantitative analysis. We found that the α1 and α4 subunits form separate populations of GABAARs and interact with distinct sets of binding proteins. We also discovered that the β3 subunit, which co-purifies with both the α1 and α4 subunits, has different levels of phosphorylation on serines 408 and 409 (S408/9) between the two receptor subtypes. To understand the role S408/9 plays in the assembly of α1- and α4-containing GABAARs, we examined the effects of S408/9A (alanine) knock-in mutation on the subunit composition of the two receptor subtypes using LC-MS/MS and quantitative analysis. We discovered that the S408/9A mutation results in the formation of novel α1α4-containing GABAARs. Moreover, in S408/9A mutants, the plasma membrane expression of the α4 subunit is increased whereas its retention in the endoplasmic reticulum is reduced. These findings suggest that S408/9 play a critical role in determining the subtype-specific assembly of GABAARs, and thus the efficacy of neuronal inhibition.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_08:58:50.578.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD036295
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Joshua Smalley
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	iodoacetamide derivatized residue
Instrument	LTQ Orbitrap Velos

Revision	Datetime	Status	ChangeLog Entry
0	2022-08-25 14:37:37	ID requested
1	2022-09-23 08:22:30	announced
⏵ 2	2023-11-14 08:58:51	announced	2023-11-14: Updated project metadata.

10.6019/PXD036295;

submitter keyword: GABA receptors, trafficking, phosphorylation, subunit composition, protein purification

Stephen James Moss
contact affiliation	Department of Neuroscience, Tufts University School of Medicine
contact email	stephen.moss@tufts.edu
lab head
Joshua Smalley
contact affiliation	Tufts University
contact email	joshua.smalley@tufts.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD036295
     1. Label: PRIDE project
     2. Name: Analyzing the mechanisms that facilitate the subtype-specific assembly of γ-aminobutyric acid type A receptors