期刊
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
卷 518, 期 1, 页码 14-18出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.bbrc.2019.07.126
关键词
Type II cGMP-dependent protein kinase; Epidermal growth factor receptor; Phosphorylation; Threonine 669; Gastric cancer cells
资金
- National Natural Science Foundation of China [31771564, 81802063, 81801168]
- Specialized Research Fund for Senior Personnel Program of Xuzhou Medical University [D2017022, D2016017]
- Natural Science Foundation of Jiangsu Province [BK20160228]
- Natural Science Fund Project of Colleges in Jiangsu Province [17KJB310001]
- Xuzhou Science and Technology planning Project [KC17096]
- Natural Science Foundation of the Jiangsu Higher Education Institution of China [18KJB320026]
Our previous study demonstrated that type II cGMP-dependent protein kinase (PKG II) inhibited epidermal growth factor (EGF) induced tyrosine phosphorylation/activation of the EGF receptor (EGFR). This paper was designed to investigate the mechanism of the inhibition of PKG II on EGFR activation. Gastric cancer cells HGC-27 and AGS were infected with an adenoviral vector encoding the cDNA of PKG II (Ad-PKG II) to overexpress PKG II and treated with 8-(4-chlorophenylthio) guanosine-30,50-cyclic monophosphate (8-pCPT-cGMP) to activate the kinase. Co-immunoprecipitation (Co-IP) and bimolecular fluorescence complementation (BiFC) assay were performed to detect the interaction between PKG II and EGFR. Western blotting, mass spectrometry (MS) and site mutagenesis were performed to detect the PKG II-specific phosphorylation site on EGFR. The results showed that in living COS-7 cells, which were infected with Ad-PKG II and treated with 8-pCPT-cGMP, there was an interaction between PKG II and EGFR. The results also showed that PKG II caused threonine 669 (T669) phosphorylation of EGFR in HGC-27 and AGS cells infected with Ad-PKG II and treated with 8-pCPT-cGMP, and then inhibited the activation of EGFR. When T669 of EGFR was mutated to alanine, the inhibitory effect of PKG II on the activation of EGFR was eradicated. These findings suggested a PKG II-specific phosphorylation site on EGFR, and might be beneficial to illuminate the anti-tumor role of PKG II. (C) 2019 Elsevier Inc. All rights reserved.
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