中文版 | English
Title

Dynamic Phosphotyrosine-Dependent Signaling Profiling in Living Cells by Two-Dimensional Proximity Proteomics

Author
Corresponding AuthorLi,Pengfei; Cai,Zongwei; Tian,Ruijun
Publication Years
2022-11-04
DOI
Source Title
ISSN
1535-3893
EISSN
1535-3907
Volume21Issue:11Pages:2727-2735
Abstract
Tyrosine phosphorylation (pTyr)-dependent signaling pathways play a vital role in various biological processes, which are spatiotemporally assembled and dynamically regulated on a minute scale by pTyr in living cells. Studying these pTyr-mediated signaling complexes is therefore challenging due to the highly dynamic nature of the protein complexes and the low abundance of pTyr. In this study, we adopted minute-resolution APEX2-based proximity labeling (PL) in living cells and Src SH2 superbinder-based pTyr peptide enrichment for simultaneously profiling these protein complexes and associated pTyr sites from the same affinity-purified sample. Upon different time courses of EGF stimulation of the living cells stably expressing APEX2-FLAG-GRB2, we constructed two-dimensional time-course curves for both interactome and tyrosine phosphoproteome. Well-annotated pTyr signaling complexes in EGFR signaling and located at the endosome were quantified with tightly correlated time-course curves for both interacting proteins and pTyr sites. Importantly, the correlated time-course curves for EGFR and endosomal HGS were well validated by targeted-parallel reaction monitoring (PRM)-MS analysis. Taking advantage of the high sensitivity of the PRM assay, the low-abundant pTyr peptide EGFR pY1110, which cannot be identified in the data-dependent acquisition (DDA) analysis, could be well quantified. Collectively, this two-dimensional proximity proteomic strategy is promising for comprehensively characterizing pTyr-mediated protein complexes with high sensitivity in living cells.
Keywords
URL[Source Record]
Indexed By
Language
English
SUSTech Authorship
First ; Corresponding
Funding Project
China State Key Basic Research Program Grants["2020YFE0202200","2021YFA1301601","2021YFA1301602"] ; National Natural Science Foundation of China["22125403","91953118","32171433","22104047","22004056"] ; Shenzhen Innovation of Science and Technology Commission["JCYJ20200109141212325","JCYJ20210324120210029","JCYJ20200109140814408"] ; Guangdong province[2019B151502050]
WOS Research Area
Biochemistry & Molecular Biology
WOS Subject
Biochemical Research Methods
WOS Accession No
WOS:000877563100001
Publisher
ESI Research Field
BIOLOGY & BIOCHEMISTRY
Scopus EID
2-s2.0-85140968982
Data Source
Scopus
Citation statistics
Cited Times [WOS]:0
Document TypeJournal Article
Identifierhttp://kc.sustech.edu.cn/handle/2SGJ60CL/411801
DepartmentDepartment of Chemistry
理学院
深圳格拉布斯研究院
Affiliation
1.Department of Chemistry,College of Science,Southern University of Science and Technology,Shenzhen,1088 Xueyuan Road,518055,China
2.Research Center for Chemical Biology and Omics Analysis,College of Science,Southern University of Science and Technology,Shenzhen,1088 Xueyuan Road,518055,China
3.Shenzhen Grubbs Institute,Southern University of Science and Technology,Shenzhen,1088 Xueyuan Road,518055,China
4.State Key Laboratory of Environmental and Biological Analysis,Department of Chemistry,Hong Kong Baptist University,Hong Kong,Kowloon Tong,999077,Hong Kong
First Author AffilicationDepartment of Chemistry;  College of Science
Corresponding Author AffilicationDepartment of Chemistry;  College of Science;  Shenzhen Grubbs Institute
First Author's First AffilicationDepartment of Chemistry;  College of Science
Recommended Citation
GB/T 7714
Kong,Qian,Ke,Mi,Weng,Yicheng,et al. Dynamic Phosphotyrosine-Dependent Signaling Profiling in Living Cells by Two-Dimensional Proximity Proteomics[J]. JOURNAL OF PROTEOME RESEARCH,2022,21(11):2727-2735.
APA
Kong,Qian.,Ke,Mi.,Weng,Yicheng.,Qin,Yunqiu.,He,An.,...&Tian,Ruijun.(2022).Dynamic Phosphotyrosine-Dependent Signaling Profiling in Living Cells by Two-Dimensional Proximity Proteomics.JOURNAL OF PROTEOME RESEARCH,21(11),2727-2735.
MLA
Kong,Qian,et al."Dynamic Phosphotyrosine-Dependent Signaling Profiling in Living Cells by Two-Dimensional Proximity Proteomics".JOURNAL OF PROTEOME RESEARCH 21.11(2022):2727-2735.
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