Original Research
Hypoxia Positively Regulates the Expression of pH-Sensing G-Protein–Coupled Receptor OGR1 (GPR68)

https://doi.org/10.1016/j.jcmgh.2016.06.003Get rights and content
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Background & Aims

A novel family of proton-sensing G-protein–coupled receptors, including ovarian cancer G-protein–coupled receptor 1 (OGR1) (GPR68) has been identified to play a role in pH homeostasis. Hypoxia is known to change tissue pH as a result of anaerobic glucose metabolism through the stabilization of hypoxia-inducible factor-1α. We investigated how hypoxia regulates the expression of OGR1 in the intestinal mucosa and associated cells.

Methods

OGR1 expression in murine tumors, human colonic tissue, and myeloid cells was determined by quantitative reverse-transcription polymerase chain reaction. The influence of hypoxia on OGR1 expression was studied in monocytes/macrophages and intestinal mucosa of inflammatory bowel disease (IBD) patients. Changes in OGR1 expression in MonoMac6 (MM6) cells under hypoxia were determined upon stimulation with tumor necrosis factor (TNF), in the presence or absence of nuclear factor-κB (NF-κB) inhibitors. To study the molecular mechanisms involved, chromatin immunoprecipitation analysis of the OGR1 promoter was performed.

Results

OGR1 expression was significantly higher in tumor tissue compared with normal murine colon tissue. Hypoxia positively regulated the expression of OGR1 in MM6 cells, mouse peritoneal macrophages, primary human intestinal macrophages, and colonic tissue from IBD patients. In MM6 cells, hypoxia-enhanced TNF-induced OGR1 expression was reversed by inhibition of NF-κB. In addition to the effect of TNF and hypoxia, OGR1 expression was increased further at low pH. Chromatin immunoprecipitation analysis showed that HIF-1α, but not NF-κB, binds to the promoter of OGR1 under hypoxia.

Conclusions

The enhancement of TNF- and hypoxia-induced OGR1 expression under low pH points to a positive feed-forward regulation of OGR1 activity in acidic conditions, and supports a role for OGR1 in the pathogenesis of IBD.

Keywords

Ovarian Cancer G-Protein–Coupled Receptor
Inflammation
Inflammatory Bowel Disease
TDAG8
GRP65

Abbreviations used in this paper

AICAR
5-aminoimidazole-4-carboxamide-1-β-4-ribofuranoside
CD
Crohn's disease
ChIP
chromatin immunoprecipitation
FCS
fetal calf serum
GPR
G-protein–coupled receptor
HIF
hypoxia-inducible factor
HV
healthy volunteer
IBD
inflammatory bowel disease
IEC
intestinal epithelial cell
IFN
interferon
IL
interleukin
MM6
MonoMac 6
mRNA
messenger RNA
NF-κB
nuclear factor-κB
OGR1
ovarian cancer G-protein–coupled receptor 1 (GPR68)
RT-qPCR
quantitative reverse-transcription polymerase chain reaction
SPARC
secreted protein acidic and rich in cysteine
TDAG8
T-cell death-associated gene 8 (GPR65)
Th
T-helper
TNF
tumor necrosis factor
UC
ulcerative colitis
WT
wild type

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Conflicts of interest The authors disclose no conflicts.

Funding This research was supported by the University of Zurich Center for Integrative Human Physiology (C.A.W. and G.R.), grants from the Swiss National Science Foundation (310030-120312 to G.R. and 31003A_155959/1 to C.A.W.), and the Swiss Inflammatory Bowel Disease Cohort (3347CO-108792). Also supported by the European Crohn's and Colitis Organisation Fellowship (J.C.-R.).

Authors share co-first authorship.