Carbon monoxide induced PPARγ SUMOylation and UCP2 block inflammatory gene expression in macrophages.

Arvand Haschemi,Martin Bilban,Markus Jeitler,Leo E Otterbein,Oswald Wagner,Beek Yoke Chin,Harald Esterbauer,Ben C B Ko

doi:10.1371/journal.pone.0026376

Arvand Haschemi, Martin Bilban + Show 6 more

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https://doi.org/10.1371/journal.pone.0026376

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Abstract

Carbon monoxide (CO) dampens pro-inflammatory responses in a peroxisome proliferator-activated receptor-γ (PPARγ) and p38 mitogen-activated protein kinase (MAPK) dependent manner. Previously, we demonstrated that CO inhibits lipopolysaccharide (LPS)-induced expression of the proinflammatory early growth response-1 (Egr-1) transcription factor in macrophages via activation of PPARγ. Here, we further characterize the molecular mechanisms by which CO modulates the activity of PPARγ and Egr-1 repression. We demonstrate that CO enhances SUMOylation of PPARγ which we find was attributed to mitochondrial ROS generation. Ectopic expression of a SUMOylation-defective PPARγ-K365R mutant partially abolished CO-mediated suppression of LPS-induced Egr-1 promoter activity. Expression of a PPARγ-K77R mutant did not impair the effect of CO. In addition to PPARγ SUMOylation, CO-activated p38 MAPK was responsible for Egr-1 repression. Blocking both CO-induced PPARγ SUMOylation and p38 activation, completely reversed the effects of CO on inflammatory gene expression. In primary macrophages isolated form C57/BL6 male mice, we identify mitochondrial ROS formation by CO as the upstream trigger for the observed effects on Egr-1 in part through uncoupling protein 2 (UCP2). Macrophages derived from bone marrow isolated from Ucp2 gene Knock-Out C57/BL6 mice (Ucp2 −/−), produced significantly less ROS with CO exposure versus wild-type macrophages. Moreover, absence of UCP2 resulted in a complete loss of CO mediated Egr-1 repression. Collectively, these results indentify p38 activation, PPARγ-SUMOylation and ROS formation via UCP2 as a cooperative system by which CO impacts the inflammatory response.

Highlights

Carbon monoxide (CO) arises physiologically in cells during the oxidative catabolism of heme by heme oxygenase (HO) enzymes [1]
CO treatment resulted in mitochondria driven ROS-dependent SUMOylation of peroxisome proliferator-activated receptor-c (PPARc) and (ii) uncoupling protein 2 (UCP2) in the mitochondria was in part responsible for ROS generation by CO
Primary Ucp22/2 macrophages confirmed a total loss of CO on ROS formation and on early growth response-1 (Egr-1) repression – compared to the partial blockade achieved by individual inhibition of p38 and PPARc-SUMOylation

Summary

Introduction

Carbon monoxide (CO) arises physiologically in cells during the oxidative catabolism of heme by heme oxygenase (HO) enzymes [1]. CO administered exogenously to cells or animals at low concentrations exerts anti-inflammatory effects in a number of preclinical models [2,3,4,5,6,7]. We demonstrated that CO suppresses LPS-induced expression of Egr-1, a key transcription factor in the generation of the inflammatory response [8] via activation of the nuclear hormone receptor PPARc [9] (reviewed in [1,10]). Since CO treatment alone does not result in generation of PPARc ligands, activity-modulating posttranslational modifications of PPARc in response to CO may occur, such as phosphorylation or SUMOylation. SUMOylation is a protein modification involving a covalent conjugation of the polypeptide SUMO to lysine residues of target proteins [14]

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