Abstract
The peptide hormone gastrin is known to play a role in differentiation, growth and apoptosis of cells in the gastric mucosa. In this study we demonstrate that gastrin induces Nuclear Receptor 4A2 (NR4A2) expression in the adenocarcinoma cell lines AR42J and AGS-GR, which both possess the gastrin/CCK2 receptor. In vivo, NR4A2 is strongly expressed in the gastrin responsive neuroendocrine ECL cells in normal mucosa, whereas gastric adenocarcinoma tissue reveals a more diffuse and variable expression in tumor cells. We show that NR4A2 is a primary early transient gastrin induced gene in adenocarcinoma cell lines, and that NR4A2 expression is negatively regulated by inducible cAMP early repressor (ICER) and zinc finger protein 36, C3H1 type-like 1 (Zfp36l1), suggesting that these gastrin regulated proteins exert a negative feedback control of NR4A2 activated responses. FRAP analyses indicate that gastrin also modifies the nucleus-cytosol shuttling of NR4A2, with more NR4A2 localized to cytoplasm upon gastrin treatment. Knock-down experiments with siRNA targeting NR4A2 increase migration of gastrin treated adenocarcinoma AGS-GR cells, while ectopically expressed NR4A2 increases apoptosis and hampers gastrin induced invasion, indicating a tumor suppressor function of NR4A2. Collectively, our results uncover a role of NR4A2 in gastric adenocarcinoma cells, and suggest that both the level and the localization of NR4A2 protein are of importance regarding the cellular responses of these cells.
Highlights
Gastrin is a gastrointestinal peptide hormone which plays a central role in regulation of gastric acid secretion [1], and in differentiation, maintenance and organization of cells/tissue in the gastric mucosa [2,3]
nuclear receptor 4A2 (NR4A2) expression is activated by gastrin Genome-wide time series experiments identified NR4A2 as a gastrin responsive gene in the pancreatic adenocarcinoma cell line AR42J (Figure 1A)
Quantitative real-time PCR confirmed that gastrin induced transient expression of NR4A2 mRNA, followed by decrease to baseline after ~6 h of stimulation; and that NR4A2 mRNA expression was sustained when protein synthesis was inhibited by CHX (Figure S1)
Summary
Gastrin is a gastrointestinal peptide hormone which plays a central role in regulation of gastric acid secretion [1], and in differentiation, maintenance and organization of cells/tissue in the gastric mucosa [2,3]. Beside its role in regulation of normal physiology, gastrin is shown to exert growth promoting impact both in normal and malignant gastrointestinal tissue. Gastrin stimulates proliferation of human gastric and pancreatic cell lines [4,5,6]. All three family members are immediate early genes induced by physiologic signals including growth factors, hormones and inflammatory cytokines [13,14], and are shown to promote cell proliferation, apoptosis and terminal differentiation in a tissue dependent manner [15,16]. NR4A2 is highly expressed in several bladder cancer cell lines and activation of the ligand-binding domain of NR4A2 was demonstrated to induce apoptotic pathways and inhibit growth of bladder cancer established in nude mice [19]. Contrary to the findings in bladder cancer cells, NR4A2 is shown to promote growth of colorectal cancer [20] and to transactivate osteopontin, a direct target of the Wnt/β-catenin pathway associated with colorectal invasion and metastasis [21]
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