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Alternative splicing of the G protein-coupled receptor superfamily in human airway smooth muscle diversifies the complement of receptors.

ABSTRACT: G protein-coupled receptors (GPCRs) are the largest signaling family in the genome, serve an expansive array of functions, and are targets for approximately 50% of current therapeutics. In many tissues, such as airway smooth muscle (ASM), complex, unexpected, or paradoxical responses to agonists/antagonists occur without known mechanisms. We hypothesized that ASM express many more GPCRs than predicted, and that these undergo substantial alternative splicing, creating a highly diversified receptor milieu. Transcript arrays were designed detecting 434 GPCRs and their predicted splice variants. In this cell type, 353 GPCRs were detected (including 111 orphans), with expression levels varying by approximately 900-fold. Receptors used for treating airway disease were expressed lower than others with similar signaling properties, indicating potentially more effective targets. A disproportionate number of Class-A peptide-group receptors, and those coupling to G(q)/(11) or G(s) (vs. G(i)), was found. Importantly, 192 GPCRs had, on average, five different expressed receptor isoforms because of splicing events, including alternative splice donors and acceptors, novel introns, intron retentions, exon(s) skips, and novel exons, with the latter two events being most prevalent. The consequences of splicing were further investigated with the leukotriene B4 receptor, known for its aberrant responsiveness in lung. We found transcript expression of three variants because of alternative donor and acceptor splice sites, representing in-frame deletions of 38 and 100 aa, with protein expression of all three isoforms. Thus, alternative splicing, subject to conditional, temporal, and cell-type regulation, is a major mechanism that diversifies the GPCR superfamily, creating local recepteromes with specialized environments.

SUBMITTER: Einstein R

PROVIDER: S-EPMC2278184 | biostudies-literature | 2008 Apr

REPOSITORIES: biostudies-literature

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Alternative splicing of the G protein-coupled receptor superfamily in human airway smooth muscle diversifies the complement of receptors.

Einstein Richard R Jordan Heather H Zhou Weiyin W Brenner Michael M Moses Esther G EG Liggett Stephen B SB

Proceedings of the National Academy of Sciences of the United States of America 20080324 13

G protein-coupled receptors (GPCRs) are the largest signaling family in the genome, serve an expansive array of functions, and are targets for approximately 50% of current therapeutics. In many tissues, such as airway smooth muscle (ASM), complex, unexpected, or paradoxical responses to agonists/antagonists occur without known mechanisms. We hypothesized that ASM express many more GPCRs than predicted, and that these undergo substantial alternative splicing, creating a highly diversified recepto ...[more]

PMID: 18362331

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Project description:?-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian central nervous system, and exerts its actions via both ionotropic (GABA(A)) and metabotropic (GABA(B)) receptors. Although the functional expression of GABA(B) receptors coupled to the G(i) protein was reported for airway smooth muscle, the role of GABA(B) receptors in airway responsiveness remains unclear. We investigated whether G(i)-coupled GABA(B) receptors cross-regulate phospholipase C (PLC), an enzyme classically regulated by G(q)-coupled receptors in human airway smooth muscle cells. Both the GABA(B)-selective agonist baclofen and the endogenous ligand GABA significantly increased the synthesis of inositol phosphate, whereas GABA(A) receptor agonists, muscimol, and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol exerted no effect. The baclofen-induced synthesis of inositol phosphate and transient increases in [Ca(2+)](i) were blocked by CGP35348 and CGP55845 (selective GABA(B) antagonists), pertussis toxin (PTX, which inactivates the G(i) protein), gallein (a G(??) signaling inhibitor), U73122 (an inhibitor of PLC-?), and xestospongin C, an inositol 1,4,5-triphosphate receptor blocker. Baclofen also potentiated the bradykinin-induced synthesis of inositol phosphate and transient increases in [Ca(2+)](i), which were blocked by CGP35348 or PTX. Moreover, baclofen potentiated the substance P-induced contraction of airway smooth muscle in isolated guinea pig tracheal rings. In conclusion, the stimulation of GABA(B) receptors in human airway smooth muscle cells rapidly mobilizes intracellular Ca(2+) stores by the synthesis of inositol phosphate via the activation of PLC-?, which is stimulated by G(??) protein liberated from G(i) proteins coupled to GABA(B) receptors. Furthermore, crosstalk between GABA(B) receptors and G(q)-coupled receptors potentiates the synthesis of inositol phosphate, transient increases in [Ca(2+)](i), and smooth muscle contraction through G(i) proteins.

Dataset Information

Alternative splicing of the G protein-coupled receptor superfamily in human airway smooth muscle diversifies the complement of receptors.

Publications

Alternative splicing of the G protein-coupled receptor superfamily in human airway smooth muscle diversifies the complement of receptors.

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