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beta-Arrestin 2 expression determines the transcriptional response to lysophosphatidic acid stimulation in murine
Diane Gesty-Palmer1, Hesham El Shewy, Trudy A Kohout
1Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Abstract:
G protein-coupled receptors often employ novel signaling mechanisms, such as transactivation of epidermal growth factor (EGF) receptors or G protein-independent signals transmitted by beta-arrestins, to control the activity of extracellular signal-regulated kinases 1 and 2 (ERK1/2). In this study we investigated the role of beta-arrestins in lysophosphatidic acid (LPA) receptor-stimulated ERK1/2 activation using fibroblast lines derived from wild type, beta-arrestin 1, beta-arrestin 2, and beta-arrestin 1/2 knock-out mice. LPA stimulation produced robust ERK1/2 phosphorylation in all four backgrounds. In cells lacking beta-arrestin 2, >80% of LPA-stimulated ERK1/2 phosphorylation was mediated by transactivated EGF receptors. In contrast, ERK1/2 activation in cells expressing beta-arrestin 2 was predominantly EGF receptor-independent. Introducing FLAG epitope-tagged beta-arrestin 2 into the beta-arrestin 1/2 null background restored EGF receptor-independent ERK1/2 activation, indicating that beta-arrestin 2 expression confers ERK1/2 activation via a distinct mechanism. To determine the contributions of beta-arrestin 2, transactivated EGF receptors, and ERK1/2 to LPA-stimulated transcriptional responses, we employed gene expression arrays containing cDNA markers for G protein-coupled receptor-mediated signaling. In the beta-arrestin 1/2 null background, 1 h of exposure to LPA significantly increased transcription of seven marker genes. Six of these responses were EGF receptor-dependent, and two required ERK1/2 activation. In beta-arrestin 2 expressing cells, three of the seven LPA-stimulated transcriptional responses observed in the beta-arrestin 1/2 null background were lost. The four residual responses were independent of EGF receptor transactivation, but all were ERK1/2-dependent. These data indicate that beta-arrestin 2 functions both to attenuate EGF receptor transactivation-dependent signaling and to promote a distinct subset of ERK1/2-mediated responses to LPA receptor activation.
Insights
Beta-arrestin 2 plays a dual role in lysophosphatidic acid (LPA) signaling, attenuating epidermal growth factor (EGF) receptor transactivation while promoting distinct extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathways.
Area of Science:
- Cellular signaling pathways
- G protein-coupled receptor (GPCR) research
- Molecular cell biology
Background:
- G protein-coupled receptors (GPCRs) utilize complex signaling mechanisms, including epidermal growth factor (EGF) receptor transactivation and beta-arrestin-mediated G protein-independent signals, to regulate extracellular signal-regulated kinases 1 and 2 (ERK1/2).
- Lysophosphatidic acid (LPA) receptors, a class of GPCRs, activate downstream signaling cascades that influence cellular processes.
Purpose of the Study:
- To investigate the specific role of beta-arrestins, particularly beta-arrestin 2, in mediating lysophosphatidic acid (LPA) receptor-stimulated extracellular signal-regulated kinase 1 and 2 (ERK1/2) activation.
- To elucidate the interplay between beta-arrestins, epidermal growth factor (EGF) receptor transactivation, and ERK1/2 signaling in response to LPA.
- To determine the contribution of these pathways to LPA-induced transcriptional changes.
Main Methods:
- Utilized fibroblast cell lines derived from wild-type, beta-arrestin 1, beta-arrestin 2, and combined beta-arrestin 1/2 knockout mice.
- Stimulated cells with LPA and assessed ERK1/2 phosphorylation and transcriptional responses using Western blotting and gene expression arrays.
- Reintroduced beta-arrestin 2 into knockout cells to confirm its specific functions.
Main Results:
- LPA robustly induced ERK1/2 phosphorylation in all cell types.
- In the absence of beta-arrestin 2, LPA-stimulated ERK1/2 activation was predominantly mediated by EGF receptor transactivation (>80%).
- Beta-arrestin 2 expression shifted LPA signaling towards EGF receptor-independent ERK1/2 activation and promoted a distinct subset of ERK1/2-dependent transcriptional responses, while attenuating EGF receptor-dependent signaling.
Conclusions:
- Beta-arrestin 2 plays a critical role in lysophosphatidic acid (LPA) receptor signaling by modulating epidermal growth factor (EGF) receptor transactivation.
- Beta-arrestin 2 promotes a unique pathway for extracellular signal-regulated kinase 1 and 2 (ERK1/2) activation that is independent of EGF receptor transactivation.
- Beta-arrestin 2 differentially regulates LPA-stimulated gene transcription, highlighting its complex role in GPCR signaling outcomes.
