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Phosphoproteomic characterization of the signaling network resulting from activation of chemokine receptor CCR2

Huang, Cheng; Foster, Simon R.; Shah, Anup D.; Kleifeld, Oded; Canals, Meritxell; Schittenhelm, Ralf B.; Stone, Martin J.

Authors

Cheng Huang

Simon R. Foster

Anup D. Shah

Oded Kleifeld

Ralf B. Schittenhelm

Martin J. Stone



Abstract

Leukocyte recruitment is a universal feature of tissue inflammation and regulated by the interactions of chemokines with their G protein-coupled receptors (GPCRs). Activation of CC chemokine receptor 2 (CCR2) by its cognate chemokine ligands, including CC chemokine ligand 2 (CCL2), plays a central role in recruitment of monocytes in several inflammatory diseases. In this study, we used phosphoproteomics to conduct an unbiased characterization of the signaling network resulting from CCL2 activation of CCR2. Using data-independent acquisition (DIA) MS analysis, we quantified both the proteome and phosphoproteome in FlpIn-HEK293T cells stably expressing CCR2 at six time points after activation with CCL2. Differential expression analysis identified 699 significantly regulated phosphorylation sites on 441 proteins. As expected, many of these proteins are known to participate in canonical signal transduction pathways and in the regulation of actin cytoskeleton dynamics, including numerous guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs). Moreover, we identified regulated phosphorylation sites in numerous proteins that function in the nucleus, including several constituents of the nuclear pore complex. The results of this study provide an unprecedented level of detail of CCR2 signaling and identify potential targets for regulation of CCR2 function.

Citation

Huang, C., Foster, S. R., Shah, A. D., Kleifeld, O., Canals, M., Schittenhelm, R. B., & Stone, M. J. (2020). Phosphoproteomic characterization of the signaling network resulting from activation of chemokine receptor CCR2. Journal of Biological Chemistry, 295, 6518-6531. https://doi.org/10.1074/jbc.ra119.012026

Journal Article Type Article
Acceptance Date Apr 2, 2020
Online Publication Date Apr 2, 2020
Publication Date May 8, 2020
Deposit Date Apr 3, 2020
Publicly Available Date Apr 3, 2020
Journal Journal of Biological Chemistry
Print ISSN 0021-9258
Electronic ISSN 1083-351X
Publisher American Society for Biochemistry and Molecular Biology
Peer Reviewed Peer Reviewed
Volume 295
Pages 6518-6531
DOI https://doi.org/10.1074/jbc.ra119.012026
Keywords Cell Biology; Biochemistry; Molecular Biology
Public URL https://nottingham-repository.worktribe.com/output/4247624
Publisher URL https://www.jbc.org/content/295/19/6518
Additional Information This research was originally published in the Journal of Biological Chemistry. Cheng Huang Simon R. Foster Anup D. Shah, Oded Kleifeld, Meritxell Canals, Ralf B. Schittenhelm, Martin J. Stone. Phosphoproteomic characterization of the signaling network resulting from activation of the chemokine receptor CCR2. J Biol Chem. 2020; 295 : p. 6518-6531. © the American Society for Biochemistry and Molecular Biology or © the Author(s).

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