Src homology 2-containing phosphotyrosine phosphatase regulates endothelin-1-induced epidermal growth factor receptor transactivation in rat renal tubular cell NRK-52E.
Epidermal growth factor (EGF) and endothelin-1 (ET-1) have been shown to be involved in proliferation and autoregeneration of renal tubular cells. This study aims to investigate the regulatory mechanism of ET-1-mediated EGF receptor (EGFR) transactivation in rat renal tubular cells (NRK-52E). Exposure of NRK-52E cells to ET-1 was found to stimulate the phosphorylation of EGFR and induce reactive oxygen species (ROS) generation. Both NAD(P)H oxidase inhibitor, diphenyliodonium (DPI) and ROS scavenger N-acetylcysteine (NAC), inhibited EGFR transactivation and extracellular signal-regulated kinase (ERK) phosphorylation caused by ET-1. In contrast, blockade of EGFR by AG1478 inhibited the phosphorylation of ERK but not ROS generation following ET-1 exposure. We found that the catalytic cysteine of Src homology 2-containing phosphotyrosine phosphatase (SHP-2) was transiently oxidized by ET-1 treatment in a modified malachite green phosphatase assay. In EGFR co-immunoprecipitation, SHP-2 was also found to interact with EGFR following ET-1 treatment. In SHP-2 knockdown NRK-52E cells, ET-1-induced EGFR transactivation was dramatically elevated and not influenced by NAC. However, GM6001 (an MMP inhibitor) and heparin binding (HB)-EGF neutralizing antibody suppressed this elevation. Our data suggest that ROS-mediated oxidation of SHP-2 is essential for HB-EGF-mediated EGFR transactivation in ET-1 signaling pathway in NRK-52E cells.
Cheng-Hsien
Chen
C
Department of Internal Medicine, Taipei Medical University-Wan Fang Hospital, No 111, Sing-Lung Road, Sec. 3, Wen-Shan District, Taipei City, 117, Taiwan.
Yung-Ho
Hsu
H
Yuh-Mou
Sue
S
Chun-Cheng
Hou
H
Horng-Mo
Lee
L
Huei-Mei
Huang
H
Tso-Hsiao
Chen
C
D000818:Animals; D002460:Cell Line; D003545:Cysteine / Q000378:metabolism; D019332:Endothelin-1 / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D007684:Kidney Tubules / Q000378:metabolism*; D010084:Oxidation-Reduction; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; D015398:Signal Transduction / Q000502:physiology; D015533:Transcriptional Activation; D018909:src Homology Domains / Q000502:physiology
D019332:Endothelin-1; D047908:Intracellular Signaling Peptides and Proteins; D017382:Reactive Oxygen Species; D004815:Epidermal Growth Factor; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D017027:Protein Tyrosine Phosphatases; C516727:Ptpn11 protein, rat; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; D003545:Cysteine