Single‐Cell Chemical Proteomics with an Activity‐Based Probe: Identification of Low‐Copy Membrane Proteins on Primary Neurons

Xu, Feng; Han, Zhiwei; Feng, Xiaojun; Chen, Linhai; Chen, Dongjuan; Yang, Zifeng; Liu, Jianfeng; Liu, Bi‐Feng

doi:10.1002/anie.201402363

Cited by 28 publications

(25 citation statements)

References 45 publications

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“…Of note, for CGP7930 there is an apparent controversy between our data where both GB1 and GB2 TMD are required to observe an agonist effect, while Binet et al, 2004 found GB2 TMD alone was sufficient. It might be due to the endogenous expression of the GB1 subunit that may exist in some cell lines including the HEK293 cells used in both studies ( Xu et al, 2014 ).…”

Section: Resultsmentioning

confidence: 99%

Allosteric ligands control the activation of a class C GPCR heterodimer by acting at the transmembrane interface

Liu

Fan

Rovira

et al. 2021

eLife

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G protein-coupled receptors (GPCRs) are among the most promising drug targets. They often form homo- and heterodimers with allosteric cross-talk between receptor entities, which contributes to fine tuning of transmembrane signaling. Specifically controlling the activity of GPCR dimers with ligands is a good approach to clarify their physiological roles and to validate them as drug targets. Here, we examined the mode of action of positive allosteric modulators (PAMs) that bind at the interface of the transmembrane domains of the heterodimeric GABAB receptor. Our site-directed mutagenesis results show that mutations of this interface impact the function of the three PAM tested. The data support the inference that they act at the active interface between both transmembrane domains, the binding site involving residues of the TM6s of the GABAB1 and the GABAB2 subunit. Importantly, the agonist activity of these PAMs involves a key region in the central core of the GABAB2 transmembrane domain, which also controls the constitutive activity of the GABAB receptor. This region corresponds to the sodium ion binding site in class A GPCRs that controls the basal state of the receptors. Overall, these data reveal the possibility of developing allosteric compounds able to specifically modulate the activity of GPCR homo- and heterodimers by acting at their transmembrane interface.

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Section: Resultsmentioning

confidence: 99%

Allosteric ligands control the activation of a class C GPCR heterodimer by acting at the transmembrane interface

Liu

Fan

Rovira

et al. 2021

eLife

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“…developed a novel single‐cell chemical proteomics strategy to profile low‐abundance membrane proteins in single cells. [ 47 ] Shi et al. built a special LIF with a fluorescent probe to quantify low copy number intracellular proteins in a single cell.…”

Section: Targeted Proteins Analysis Approachesmentioning

confidence: 99%

“…Xu et al developed a novel single-cell chemical proteomics strategy to profile low-abundance membrane proteins in single cells. [47] Shi et al built a special LIF with a fluorescent probe to quantify low copy number intracellular proteins in a single cell. [48] However, due to the vulnerability to the adsorption of proteins on the capillary wall, the detection stability is always a big challenge to the capillary-based separation method, bringing difficulties in practical application.…”

Section: Capillary Electrophoresis-laser-induced Fluorescence Detectionmentioning

confidence: 99%

The Intriguing Landscape of Single‐Cell Protein Analysis

Xie

2022

Advanced Science

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Profiling protein expression at single‐cell resolution is essential for fundamental biological research (such as cell differentiation and tumor microenvironmental examination) and clinical precision medicine where only a limited number of primary cells are permitted. With the recent advances in engineering, chemistry, and biology, single‐cell protein analysis methods are developed rapidly, which enable high‐throughput and multiplexed protein measurements in thousands of individual cells. In combination with single cell RNA sequencing and mass spectrometry, single‐cell multi‐omics analysis can simultaneously measure multiple modalities including mRNAs, proteins, and metabolites in single cells, and obtain a more comprehensive exploration of cellular signaling processes, such as DNA modifications, chromatin accessibility, protein abundance, and gene perturbation. Here, the recent progress and applications of single‐cell protein analysis technologies in the last decade are summarized. Current limitations, challenges, and possible future directions in this field are also discussed.

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“…A chemical cytometry-based on microfluidics capillary electrophoresis technology was used for single-cell chemical proteomics (SCCP) to recognize proteins present on the individual primary neuron membrane. A functional single-neuron was isolated from a mouse and injected into a capillary for generating capillary electrophoresis-laser-induced fluorescence to study receptor protein, labeled by activity-based probes [ 90 ]. The electropherogram results conclude the heterogeneous expression of GB1 probably owing to the presence of two important G-protein (GB1) variant splices (GB1a and GB1b, in CNS) or precise post-translational alterations.…”

Section: Microfluidics-based Single-neuron Omicsmentioning

confidence: 99%