Oxidative Stress Activates the Human Histidine Decarboxylase Promoter in AGS Gastric Cancer Cells

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179

149

Enhanced VEGF-A (vascular endothelial growth factor A) gene expression is associated with increased tumor growth and metastatic spread of solid malignancies including gastric cancer. Oxidative stress has been linked to tumor-associated neoangiogenesis; underlying mechanisms, however, remained poorly understood. Therefore, we studied the effect of oxidative stress on VEGF-A gene expression in gastric cancer cells. Oxidative stress generated by H 2 O 2 application potently stimulated VEGF-A protein and mRNA levels as determined by enzyme-linked immunosorbent assay and real-time PCR techniques, respectively, and elevated the activity of a transfected (؊2018) VEGF-A promoter reporter gene construct in a time-and dose-dependent manner (4 -8-fold). These effects were abolished by the antioxidant N-acetylcysteine, demonstrating specificity of oxidative stress responses. Functional 5 deletion analysis mapped the oxidative stress response element of the human VEGF-A promoter to the sequence ؊88/؊50, and a single copy of this element was sufficient to confer basal promoter activity as well as oxidative stress responsiveness to a heterologous promoter system. Combination of EMSA studies, Sp1/Sp3 overexpression experiments in Drosophila SL-2 cells, and systematic promoter mutagenesis identified enhanced Sp1 and Sp3 binding to two GC-boxes at ؊73/؊66 and ؊58/؊52 as the core mechanism of oxidative stress-triggered VEGF-A transactivation. Additionally, in Gal4-Sp1/-Sp3-Gal4-luciferase assays, oxidative stress increased Sp1 but not Sp3 transactivating capacity, indicating additional mechanism(s) of VEGF-A gene regulation. Signaling studies identified a cascade comprising Ras 3 Raf 3 MEK1 3 ERK1/2 as the main pathway mediating oxidative stress-stimulated VEGF-A transcription. This study for the first time delineates the mechanisms underlying regulation of VEGF-A gene transcription by oxidative stress and thereby further elucidates potential pathways underlying redox control of neoangiogenesis.

Section: Discussionsupporting

confidence: 72%

Section: Discussionmentioning

confidence: 58%

Oxidative Stress Regulates Vascular Endothelial Growth Factor-A Gene Transcription through Sp1- and Sp3-dependent Activation of Two Proximal GC-rich Promoter Elements

Schäfer

Cramer

Suske

et al. 2003

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179

149

“…In addition to its central role in regulation of gastric acid secretion, increased HDC gene expression has also been found to be associated with gastric inflammation and development of gastroduodenal ulceration (31)(32)(33)(34)(35). Furthermore, we found that oxidative stress, which is commonly increased in ulcerative and inflammatory diseases affecting the gastric mucosa, is capable of transactivating the hHDC promoter in vitro (36). Aside from its ulcerogenic potential, histamine has also been shown to possess immunomodulatory properties in the context of mucosal inflammations, contributes to the healing of ulcerative lesions of the gastric and intestinal mucosa, and has also been shown to stimulate the growth of gastric epithelial cells (37)(38)(39)(40)(41).…”

mentioning

confidence: 64%

“…Although some overlap between both pathways has been described (46), the JNK cascade activates primarily transcription factors involved in the "stress response" of eukaryotic cells, whereas the ERK pathway has been linked to genes involved in cellular proliferation and differentiation (46 -48). Recently we demonstrated that ERK-related signaling cascades also play a central role in the transmission of the effects of gastrin and oxidative stress on the hHDC promoter in gastric cancer cells, whereas the JNK pathway is not involved in hHDC gene regulation (36,49). Therefore, it is highly likely that the ERK cascade represents a potential target signaling route through which activators of hHDC gene transcription exert their transactivating effect on the hHDC promoter.…”

mentioning

confidence: 99%

Helicobacter pylori Activates the Histidine Decarboxylase Promoter through a Mitogen-activated Protein Kinase Pathway Independent of Pathogenicity Island-encoded Virulence Factors

Weßler¹,

Höcker²,

Fischer³

et al. 2000

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Helicobacter pylori infection of the gastric mucosa is accompanied by an activated histamine metabolism. Histamine plays a central role in the regulation of gastric acid secretion and is involved in the pathogenesis of gastroduodenal ulcerations. Histidine decarboxylase (HDC) is the rate-limiting enzyme for histamine production, and its activity is regulated through transcriptional mechanisms. The present study investigated the effect of H. pylori infection on the transcriptional activity of the human HDC (hHDC) promoter in a gastric epithelial cell line (AGS) and analyzed the underlying molecular mechanisms. Our studies demonstrate that H. pylori infection potently transactivated the hHDC promoter. The H. pylori-responsive element of the hHDC gene was mapped to the sequence ؉1 to ؉27 base pairs, which shows no homology to known cis-acting elements and also functions as a gastrin-responsive element. H. pylori regulates the activity of this element via a Raf-1/ MEK/ERK pathway, which was activated in a Ras-independent manner. Furthermore, we found that H. pyloriinduced transactivation of the hHDC promoter was independent of the cag pathogenicity island and the vacuolating cytotoxin A gene and therefore may be exerted through (a) new virulence factor(s). A better understanding of H. pylori-directed hHDC transcription can provide novel insights into the molecular mechanisms of H. pylori-dependent gene regulation in gastric epithelial cells and may lead to new therapeutic approaches.

“…HDC promoter activity is up-regulated by several different stimuli, including gastrin (8), phorbol ester phorbol 12-myristate 13-acetate (8 -11), oxidative stress (12), thrombopointin (13), and Helicobacter pylori infection (14,15). Whereas not all of the cis-acting DNA elements or the transcriptional factors involved in regulation of HDC transcription have been identified, three GC-rich gastrin responsive elements located downstream of the transcription initiation site have been characterized in the human HDC promoter region (16,17).…”

mentioning

confidence: 99%

Kruppel-like Factor 4 (KLF4) Represses Histidine Decarboxylase Gene Expression through an Upstream Sp1 Site and Downstream Gastrin Responsive Elements

Liu

Langlois

et al. 2004

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Histidine decarboxylase (HDC) is the enzyme that catalyzes the conversion of histidine to histamine, a bioamine that plays an important role in allergic responses, inflammation, neurotransmission, and gastric acid secretion. Previously, we demonstrated that gastrin activates HDC promoter activity in a gastric cancer (AGS-E) cell line through three overlapping downstream promoter elements. In the current study, we used the yeast one-hybrid strategy to identify nuclear factors that bind to these three elements. Among eight positives from the one-hybrid screen, we identified Kruppel-like factor 4 (KLF4) (previously known as gut-enriched Kruppel-like factor (GKLF)) as one factor that binds to the gastrin responsive elements in the HDC promoter. Electrophoretic mobility shift assays confirmed that KLF4 is able to bind all three gastrin responsive elements. In addition, transient cotransfection experiments showed that overexpression of KLF4 dose dependently and specifically inhibited HDC promoter activity. Regulation of HDC transcription by KLF4 was confirmed by changes in the endogenous HDC messenger RNA by KLF4 small interfering RNA and KLF4 overexpression. We further showed that KLF4 inhibits HDC promoter activity by competing with Sp1 at the upstream GC box and also independently by binding the three downstream gastrin responsive elements. Taken together, these results indicate that KLF4 can act to repress HDC gene expression by Sp1-dependent and -independent mechanisms.Histamine is a bioamine that plays an important role in many physiological processes, including allergy, inflammation, neurotransmission, and gastric acid secretion (1-3). Histidine decarboxylase (HDC) 1 is the single enzyme that converts histidine to histamine (4). HDC is expressed in many different cell types, including mast cells, skin cells, platelets, and basophils. However, in the adult mammals, HDC is highly expressed in enterochromaffin-like cells, where the HDC activity is tightly regulated by a gut peptide hormone, gastrin (5). HDC regulation occurs at both the transcriptional and post-translational levels, the latter by proteolytic processing through the ubiquitin-proteasome pathway (6, 7).HDC promoter activity is up-regulated by several different stimuli, including gastrin (8), phorbol ester phorbol 12-myristate 13-acetate (8 -11), oxidative stress (12), thrombopointin (13), and Helicobacter pylori infection (14, 15). Whereas not all of the cis-acting DNA elements or the transcriptional factors involved in regulation of HDC transcription have been identified, three GC-rich gastrin responsive elements located downstream of the transcription initiation site have been characterized in the human HDC promoter region (16,17). Through the use of Southwestern blot and UV cross-linking, the sizes of the three gastrin responsive element-binding factors have previously been assessed at 53, 33, and 110 kDa of apparent molecular mass, respectively (16,17). Recently, a neural peptide pituitary adenylate cyclase-activating polypeptide has been reporte...