Quantitative Multiple-Reaction Monitoring Proteomic Analysis of Gβ and Gγ Subunits in C57Bl6/J Brain Synaptosomes,Biochemistry

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Quantitative Multiple-Reaction Monitoring Proteomic Analysis of Gβ and Gγ Subunits in C57Bl6/J Brain Synaptosomes
Biochemistry ( IF 2.9 ) Pub Date : 2017-09-21 00:00:00 , DOI: 10.1021/acs.biochem.7b00433
Yun Young Yim ₁ , W. Hayes McDonald ₂ , Karren Hyde ₁ , Osvaldo Cruz-Rodríguez , John J. G. Tesmer , Heidi E. Hamm ₁

Affiliation

Gβγ dimers are one of the essential signaling units of activated G protein-coupled receptors (GPCRs). There are five Gβ and 12 Gγ subunits in humans; numerous studies have demonstrated that different Gβ and Gγ subunits selectively interact to form unique Gβγ dimers, which in turn may target specific receptors and effectors. Perturbation of Gβγ signaling can lead to impaired physiological responses. Moreover, previous targeted multiple-reaction monitoring (MRM) studies of Gβ and Gγ subunits have shown distinct regional and subcellular localization patterns in four brain regions. Nevertheless, no studies have quantified or compared their individual protein levels. In this study, we have developed a quantitative MRM method not only to quantify but also to compare the protein abundance of neuronal Gβ and Gγ subunits. In whole and fractionated crude synaptosomes, we were able to identify the most abundant neuronal Gβ and Gγ subunits and their subcellular localizations. For example, Gβ₁ was mostly localized at the membrane while Gβ₂ was evenly distributed throughout synaptosomal fractions. The protein expression levels and subcellular localizations of Gβ and Gγ subunits may affect the Gβγ dimerization and Gβγ–effector interactions. This study offers not only a new tool for quantifying and comparing Gβ and Gγ subunits but also new insights into the in vivo distribution of Gβ and Gγ subunits, and Gβγ dimer assembly in normal brain function.

中文翻译：

C57Bl6 / J脑突触小体中Gβ和Gγ亚基的定量多反应监测蛋白质组学分析

Gβγ二聚体是激活的G蛋白偶联受体（GPCR）的重要信号单元之一。人中有五个Gβ和12个Gγ亚基。大量研究表明，不同的Gβ和Gγ亚基选择性相互作用形成独特的Gβγ二聚体，进而可能靶向特定的受体和效应子。Gβγ信号的扰动可能导致生理反应受损。此外，先前针对Gβ和Gγ亚基的靶向多反应监测（MRM）研究表明，在四个大脑区域中存在明显的区域和亚细胞定位模式。然而，尚无研究量化或比较其各自的蛋白质水平。在这项研究中，我们开发了一种定量MRM方法，不仅可以定量，而且可以比较神经元Gβ和Gγ亚基的蛋白质丰度。在完整和分级分离的粗突触体中，我们能够鉴定出最丰富的神经元Gβ和Gγ亚基及其亚细胞定位。例如，Gβ₁在将膜大多局部而Gβ ₂被均匀分布在整个的突触体的级分分布。Gβ和Gγ亚基的蛋白表达水平和亚细胞定位可能影响Gβγ二聚化和Gβγ-效应子相互作用。这项研究不仅为定量和比较Gβ和Gγ亚基提供了新的工具，而且为Gβ和Gγ亚基在体内的分布以及Gβγ二聚体在正常脑功能中的体内分布提供了新见解。

更新日期：2017-09-21

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