Roles of Pyk2 in signal transduction after gonadotropin‐releasing hormone receptor stimulation

• Published in: Journal of cellular physiology Vol. 236; no. 4; pp. 3033 - 3043
• Main Authors: Okitsu‐Sakurayama, Shiho, Higa‐Nakamine, Sayomi, Torihara, Hidetsugu, Higashiyama, Shigeki, Yamamoto, Hideyuki
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.04.2021
• Subjects: Antibodies; Anticoagulants; Epidermal growth factor; Epidermal growth factor receptors; Extracellular signal-regulated kinase; Flags; Gonadotropin-releasing hormone; Gonadotropins; GRB2 protein; Growth factors; Heparin; heparin‐binding EGF‐like growth factor; Hypothalamus; Kinases; mitogen‐activated protein kinase 1; mitogen‐activated protein kinase 3; Neurons; Phenylalanine; Pituitary (anterior); Proline; Protein-tyrosine kinase; Proteins; PTK2B protein; Raf protein; Receptors; Signal transduction; siRNA; Tyrosine; gonadotropin-releasing hormone; heparin-binding EGF-like growth factor; PTK2B protein; mitogen-activated protein kinase 1; GRB2 protein; mitogen-activated protein kinase 3
• ISSN: 0021-9541; 1097-4652
• DOI: 10.1002/jcp.30077

The receptor for gonadotropin‐releasing hormone (GnRH) is highly expressed in hypothalamic neurons. It has been reported that GnRH treatment of cultured GnRH neurons (GT1‐7 cells) activated proline‐rich tyrosine kinase 2 (Pyk2), and Pyk2 was involved in the activation of extracellular signal‐regulated protein kinase 1 (ERK1) and ERK2 (ERK1/2). In the present study, we first examined the possibility that GnRH treatment might activate epidermal growth factor receptor (EGFR). We found that activation of EGFR after GnRH treatment for 5 min was much less than after EGF or heparin‐binding EGF treatment. Next, we examined whether or not Pyk2 bound to growth factor receptor‐binding protein 2 (Grb2). We overexpressed FLAG‐fused Pyk2 in GT1‐7 cells, and immunoprecipitated Pyk2 using an anti‐FLAG antibody. The binding of Pyk2 to Grb2 was detected only after GnRH treatment. In contrast, a site‐directed mutant of Pyk2 wherein tyrosine 881 was mutated to phenylalanine did not bind to Grb2. Studies with small interfering RNA and inhibitors indicated that the activation of Grb2/Ras/Raf/MEK was a major pathway to ERK1/2 activation after the short‐term treatment of GT1‐7 cells with GnRH. Activated Pyk2 activates ERK1/2 through the Grb2/Ras/Raf/MEK pathway. GnRH long term treatment induces the shedding of proHB‐EGF.