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Identification, cloning, and functional characterization of a murine lipoxin A4 receptor homologue gene
Michael W Vaughn1, Rita J Proske, David L Haviland
1Institute of Molecular Medicine for the Prevention of Human Diseases, Research Center for Immunology and Autoimmune Diseases, University of Texas-Houston Health Science Center, Houston, TX 77030, USA.
Abstract:
To identify additional members of the murine N-formyl-Met-Leu-Phe peptide receptor family (fMLF-R), a mouse macrophage cDNA library was screened using the open reading frame of murine N-formyl peptide receptor. Four individual hybridizing cDNA clones were maintained through tertiary screening. One cDNA clone was a truncated, polyadenylated version of the previously described murine-fMLF-R. The other three cDNA clones varied in length, but contained identical open reading frame sequences. One clone, 8C10, was selected for further study and shared 70% sequence identity with murine-fMLF-R and 89% sequence identity with murine lipoxin A4 receptor cDNA. When placed into the pcDNA-3 expression vector and cotransfected with Galpha16 cDNA into COS-1 cells, 8C10 cDNA induced the production of inositol-1,4,5-triphosphate when concentrations of 1-1600 nM lipoxin A4 (LXA4) were tested as ligands. Northern blot analysis of murine organs indicated that the 8C10 message is present in lung, spleen, and adipose tissue. Moreover, mice treated with LPS demonstrated increased expression of 8C10 message in spleen and adipose tissue, while showing a slight reduction in lung. We have also characterized the 8C10 structural gene from a 129Sv/J genomic library and have determined its size to be >6.1 kb in length and comprised of two exons separated by a 4.8-kb intron. Collectively, these data indicate that this homologue receptor is closely related to the murine LXA4 receptor and functionally responds to LXA4 as a ligand.
Insights
Researchers identified a novel murine receptor, 8C10, closely related to the lipoxin A4 receptor. This receptor is activated by lipoxin A4 (LXA4) and its expression is modulated by lipopolysaccharide (LPS) in mice.
Area of Science:
- Immunology and Pharmacology
- Molecular Biology and Genetics
Background:
- The N-formyl-Met-Leu-Phe peptide receptor (fMLF-R) family plays a crucial role in inflammatory responses.
- Identifying novel receptors within this family is essential for understanding immune signaling pathways.
Purpose of the Study:
- To discover new members of the murine N-formyl-Met-Leu-Phe peptide receptor family.
- To characterize the function and expression of a newly identified receptor homologue.
Main Methods:
- Screening of a mouse macrophage cDNA library using the murine N-formyl peptide receptor open reading frame.
- Sequence analysis, gene cloning, and functional assays (inositol phosphate production) in COS-1 cells.
- Northern blot analysis of murine organs and gene characterization from a genomic library.
Main Results:
- A novel cDNA clone, 8C10, was identified, sharing high sequence identity with the murine lipoxin A4 receptor (89%).
- The 8C10 receptor functionally responded to lipoxin A4 (LXA4) in a dose-dependent manner, inducing inositol phosphate production.
- 8C10 mRNA was detected in lung, spleen, and adipose tissue, with altered expression in spleen and adipose tissue following LPS treatment.
Conclusions:
- The 8C10 receptor is a functional homologue of the murine LXA4 receptor, activated by LXA4.
- Its expression is tissue-specific and dynamically regulated by inflammatory stimuli like LPS.
- This finding expands the known repertoire of LXA4 receptors and their roles in immune regulation.