Abstract
The cysteinyl-leukotrienes (cysLTs) LTC4, LTD4, and LTE4, are involved in a variety of inflammatory diseases, including asthma, and act on at least two distinct receptors, CysLT1 and CysLT2. Specific antagonists of CysLT1 are currently used to control bronchoconstriction and inflammation in asthmatic patients. The potential role of CysLT2 in asthma remains poorly understood. A polymorphism in the CysLT2 gene, resulting in a single amino acid substitution (M201V), was found to be associated with asthma in three separate population studies. Here, we investigated whether the M201V mutation affected the affinity of CysLT2 for its natural ligands and its signaling efficiency. Human embryonic kidney 293 cells were stably transfected with either wild-type (wt) or mutant (M201V) CysLT2. Affinity of the M201V receptor for LTC4 was reduced by 50%, whereas affinity for LTD4 was essentially lost. LTC4-induced production of inositol phosphates (IPs) in M201V-expressing cells was significantly decreased at suboptimal concentrations of the ligand, but no difference was observed at high concentrations. In contrast, LTD4-induced IP production was 10- to 100-fold less in M201V- than in wt-expressing cells. Similar results were also observed with the transactivation of the interleukin-8 promoter induced by LTC4 or LTD4. Moreover, in contrast to wt-expressing cells, phosphorylation of nuclear factor κB p65 was absent in LTD4-stimulated M201V-expressing cells. Likewise, phosphorylation of c-Jun N-terminal kinase was not induced in LTD4-stimulated M201V cells, whereas activation of extracellular response kinase and p38 was maintained, at least at higher LTD4 concentrations. Our results indicate that the M201V polymorphism drastically affects CysLT2 responses to LTD4 and less to LTC4.
Footnotes
This work was supported by the Canadian Institutes of Health Research [Grant MOP 82716] (to J.S. and M.R.-P.); a Canada Research Chair in Inflammation (to M.R.-P); and Centre de Recherche Clinique Étienne-Le Bel, funded by Fonds de la Recherche en Santé du Québec (to M.R.-P. and J.S.)
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.110.172411.
-
ABBREVIATIONS:
- cysLT
- cysteinyl-leukotriene
- CysLT
- cysLT receptor
- LT
- leukotriene
- AP-1
- activating protein 1
- ECL
- extracellular loop
- ERK
- extracellular response kinase
- HEK
- human embryonic kidney
- IL
- interleukin
- IP
- inositol phosphate
- JNK
- c-Jun N-terminal kinase
- MAPK
- mitogen-activated protein kinase
- NF-κB
- nuclear factor κB
- TM
- transmembrane helix
- wt
- wild type
- ANOVA
- analysis of variance
- GPCR
- G protein-coupled receptor
- DMEM
- Dulbecco's modified Eagle's medium
- BAY u9773
- 4-[[(1R,2E,4E,6Z,9Z)-1-[(1S)-4-carboxy-1-hydroxybutyl]-2,4,6,9-pentadecatetraenyl]thio]-benzoic acid.
- Received July 1, 2010.
- Accepted October 4, 2010.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|