Proteomics analysis of G protein-coupled receptor kinase 4-inhibited cellular growth of HEK293 cells

Yunxiu Luo; Xishi Huang; Jing Yang; Lanzhen Huang; Ruirui Li; Qunyin Wu; Xiaoshan Jiang

doi:10.1016/j.jprot.2019.103445

Proteomics analysis of G protein-coupled receptor kinase 4-inhibited cellular growth of HEK293 cells

J Proteomics. 2019 Sep 15:207:103445. doi: 10.1016/j.jprot.2019.103445. Epub 2019 Jul 23.

Authors

Yunxiu Luo¹, Xishi Huang², Jing Yang³, Lanzhen Huang³, Ruirui Li², Qunyin Wu⁴, Xiaoshan Jiang⁵

Affiliations

¹ Xiangya Hospital, Central South University, Changsha, Hunan 410008, China; Cell Signaling Laboratory, China.
² Cell Signaling Laboratory, China.
³ Center for Science Research, China.
⁴ Department of Genetics and Cell Biology, Guilin Medical University, Guilin, Guangxi 541004, China.
⁵ Cell Signaling Laboratory, China; Center for Science Research, China; Department of Oncology at the 2nd Affiliated Hospital, Guilin Medical University, Guilin, Guangxi 541004, China.. Electronic address: jiangxs@glmc.edu.cn.

PMID: 31349021
DOI: 10.1016/j.jprot.2019.103445

Abstract

G protein-coupled receptor kinases (GRKs) are involved in a wide range of cellular physiology and pathological activities by specifically phosphorylating activated G protein-coupled receptors (GPCRs) to terminate GPCR signaling, or through regulating non-GPCR substrates. We recently reported that overexpression of GRK4 halts cell proliferation and induces cellular senescent phenotype in HEK293 cells. In this study, a quantitative proteomic assay was performed to analyze the protein profiles between HEK293 cells expressing and not expressing GRK4. Results revealed 39 upregulated and 59 downregulated differently expressed proteins (DEPs) in a total of 4124 identified proteins. Gene ontology (GO) annotation and functional enrichment revealed that the DEPs were related to metabolic processes regulated by the binding of these RNA/proteins under the biological processes. The Kyoto Encyclopedia of Gene and Genomes (KEGG) analysis showed pathways of cell development, division, proliferation, apoptosis, aging, autophagy, cell death and cell cycle progression are involved in. Immunoblotting validation of expression of six key target proteins, CALM1, STAT3, CDK1, CDK6, TOP2A, and GRK4, which speculatively maintain abnormal activity in the above pathways, was consistent with the results of proteomics analysis. Lastly, a biological phenotype assay confirmed that GRK4 promoted HEK293 cell growth blockage and G1/0 arrest. Taken together, this study identified some novel molecules that involve in GRK4 signaling and provided valuable information for further studying the mechanisms underlying GRK4-induced proliferative inhibition. SIGNIFICANCE: A quantitative proteomic assay was performed in HEK293 cells expressing and not expressing GRK4 39 upregulated and 59 downregulated differently expressed proteins (DEPs)were identified. DEPs involved in pathways of cell development, division, proliferation, apoptosis, aging, autophagy, cell death and cell cycle progression. Biological phenotype assay confirmed that GRK4 prompted HEK293 cell growth blockage and G1/0 arrest.

Keywords: Cell growth; G protein-coupled receptor kinase 4; Quantitative proteomics.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

G-Protein-Coupled Receptor Kinase 4 / metabolism*
G1 Phase Cell Cycle Checkpoints*
Gene Expression Regulation*
HEK293 Cells
Humans
Proteomics*
Resting Phase, Cell Cycle*
Signal Transduction*

Substances

G-Protein-Coupled Receptor Kinase 4
GRK4 protein, human