Helicobacter pylori-induced cell death is counteracted by NF-κB-mediated transcription of DARPP-32

Shoumin Zhu, Mohammed Soutto, Zheng Chen, Dunfa Peng, Judith Romero-Gallo, Uma S. Krishna, Abbes Belkhiri, M. Kay Washington, Richard Peek, Wael El-Rifai

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13 Scopus citations

Abstract

Objective DARPP-32 is a frequently amplified and overexpressed gene that promotes several oncogenic functions in gastric cancer. Herein, we investigated the relationship between Helicobacter pylori infection, proinflammatory NF-κB activation and regulation of DARPP-32. Design The study used in vivo and in vitro experiments. Luciferase reporter, quantitative real-time PCR, immunoblot, chromatin immunoprecipitation (ChIP), cell viability, H. pylori infection, tissue microarrays and immunohistochemical assays were used. Results Our results indicated that H. pylori infection increased the DARPP-32 mRNA and protein levels in gastric cancer cell lines and gastric mucosa of mice. H. pylori infection increased the activity of NF-κB reporter and p-NF-κB (S536) protein level in vitro and in vivo. To investigate the transcriptional regulation of DARPP-32, we cloned a 3019 bp of the DARPP-32 promoter into the luciferase reporter ( pGL3-Luc). Both H. pylori infection and tumour necrosis factor-α treatment induced DARPP-32 reporter activity ( p<0.01). Using deletion constructs of DARPP-32 promoter and ChIP assay, we demonstrated that the sequence -996 to -1008 bp containing putative NF-κB-binding sites is the most active region. The induction of DARPP-32 expression by H. pylori infection counteracted H. pyloriinduced cell death through activation of serine/threoninespecific protein kinase (AKT), as determined by ATP-Glo and clonogenic survival assays. Immunohistochemistry analysis demonstrated a significant positive correlation between NF-κB and DARPP-32 expression levels in gastric cancer tissues (r2=0.43, p<0.01). Conclusions Given the high frequency of DARPP-32 overexpression and its prosurvival oncogenic functions, the induction of DARPP-32 expression following H. pylori infection and activation of NF-κB provides a link between infection, inflammation and gastric tumourigenesis.

Original languageEnglish (US)
Pages (from-to)802-812
Number of pages11
JournalGut
Volume66
Issue number5
DOIs
StatePublished - Sep 2 2016
Externally publishedYes

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ASJC Scopus subject areas

  • Gastroenterology

Cite this

Zhu, S., Soutto, M., Chen, Z., Peng, D., Romero-Gallo, J., Krishna, U. S., Belkhiri, A., Washington, M. K., Peek, R., & El-Rifai, W. (2016). Helicobacter pylori-induced cell death is counteracted by NF-κB-mediated transcription of DARPP-32. Gut, 66(5), 802-812. https://doi.org/10.1136/gutjnl-2016-312141