Affiliation:
1. Departments of Physiology and Medicine, Medical College of Virginia Campus, Virginia Commonwealth University, Richmond, VA 23298, U.S.A.
Abstract
Initial Ca2+-dependent contraction of the intestinal smooth muscle mediated by Gq-coupled receptors is attenuated by RGS4 (regulator of G-protein signalling 4). Treatment of colonic muscle cells with IL-1β (interleukin-1β) inhibits acetylcholine-stimulated initial contraction through increasing the expression of RGS4. NF-κB (nuclear factor κB) signalling is the dominant pathway activated by IL-1β. In the present study we show that RGS4 is a new target gene regulated by IL-1β/NF-κB signalling. Exposure of cultured rabbit colonic muscle cells to IL-1β induced a rapid increase in RGS4 mRNA expression, which was abolished by pretreatment with a transcription inhibitor, actinomycin D, implying a transcription-dependent mechanism. Existence of the canonical IKK2 [IκB (inhibitor of NF-κB) kinase 2]/IκBα pathway of NF-κB activation induced by IL-1β in rabbit colonic muscle cells was validated with multiple approaches, including the induction of reporter luciferase activity and endogenous NF-κB-target gene expression, NF-κB-DNA binding activity, p65 nuclear translocation, IκBα degradation and the phosphorylation of IKK2 at Ser177/181 and p65 at Ser536. RGS4 up-regulation by IL-1β was blocked by selective inhibitors of IKK2, IκBα or NF-κB activation, by effective siRNA (small interfering RNA) of IKK2, and in cells expressing either the kinase-inactive IKK2 mutant (K44A) or the phosphorylation-deficient IκBα mutant (S32A/S36A). An IKK2-specific inhibitor or effective siRNA prevented IL-1β-induced inhibition of acetylcholine-stimulated PLC-β (phopsholipase C-β) activation. These results suggest that the canonical IKK2/IκBα pathway of NF-κB activation mediates the up-regulation of RGS4 expression in response to IL-1β and contributes to the inhibitory effect of IL-1β on acetylcholine-stimulated PLC-β-dependent initial contraction in rabbit colonic smooth muscle.
Subject
Cell Biology,Molecular Biology,Biochemistry
Cited by
38 articles.
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