MAGI1 as a link between endothelial activation and ER stress drives atherosclerosis
Jun-ichi Abe, … , Scott E. Evans, Nhat-Tu Le
Jun-ichi Abe, … , Scott E. Evans, Nhat-Tu Le
Published April 4, 2019
Citation Information: JCI Insight. 2019;4(7):e125570. https://doi.org/10.1172/jci.insight.125570.
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Research Article Cardiology Cell biology

MAGI1 as a link between endothelial activation and ER stress drives atherosclerosis

Abstract

The possible association between the membrane-associated guanylate kinase with inverted domain structure-1 (MAGI1) and inflammation has been suggested, but the molecular mechanisms underlying this link, especially during atherogenesis, remain unclear. In endothelial cells (ECs) exposed to disturbed flow (d-flow), p90 ribosomal S6 kinase (p90RSK) bound to MAGI1, causing MAGI1-S741 phosphorylation and sentrin/SUMO-specific protease 2 T368 phosphorylation-mediated MAGI1-K931 deSUMOylation. MAGI1-S741 phosphorylation upregulated EC activation via activating Rap1. MAGI1-K931 deSUMOylation induced both nuclear translocation of p90RSK-MAGI1 and ATF-6-MAGI1 complexes, which accelerated EC activation and apoptosis, respectively. Microarray screening revealed key roles for MAGI1 in the endoplasmic reticulum (ER) stress response. In this context, MAGI1 associated with activating transcription factor 6 (ATF-6). MAGI1 expression was upregulated in ECs and macrophages found in atherosclerotic-prone regions of mouse aortas as well as in the colonic epithelia and ECs of patients with inflammatory bowel disease. Further, reduced MAGI1 expression in Magi1–/+ mice inhibited d-flow–induced atherogenesis. In sum, EC activation and ER stress–mediated apoptosis are regulated in concert by two different types of MAGI1 posttranslational modifications, elucidating attractive drug targets for chronic inflammatory disease, particularly atherosclerosis.

Authors

Jun-ichi Abe, Kyung Ae Ko, Sivareddy Kotla, Yin Wang, Jesus Paez-Mayorga, Ik Jae Shin, Masaki Imanishi, Hang Thi Vu, Yunting Tao, Miguel M. Leiva-Juarez, Tamlyn N. Thomas, Jan L. Medina, Jong Hak Won, Yuka Fujii, Carolyn J. Giancursio, Elena McBeath, Ji-Hyun Shin, Liliana Guzman, Rei J. Abe, Jack Taunton, Naoki Mochizuki, William Faubion, John P. Cooke, Keigi Fujiwara, Scott E. Evans, Nhat-Tu Le

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Figure 5

MAGI1 deSUMOylation associated with EC activation.

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MAGI1 deSUMOylation associated with EC activation. (A and B) HUVECs trea...
(A and B) HUVECs treated with FMK-MEA (10 μM, 30 minutes) were exposed to d-flow, and the level of SUMOylated MAGI1 was determined (first row above the dotted line). The membrane was then immunoblotted with an anti-MAGI1 antibody (A, Sigma, rabbit polyclonal antibody [M5691], and B, Santa Cruz, mouse monoclonal antibody [SC-100326]) (second row above the dotted line), indicating pull-down of equal amounts of MAGI1. The levels of SUMO2/3 and tubulin expression in cell lysates are shown in the rows below the dotted line. SUMOylated MAGI1 levels were quantified by densitometry of immunoblots from 3 independent experiments and are plotted (B) (n = 3). Data represent mean ± SEM. **P < 0.01. Cont, control. (C) HUVECs transduced with Ad-SENP2-WT or -T368A were exposed to d-flow, and the level of SUMOylated MAGI1, MAGI1, SENP2, and SUMO2/3 expression was determined as in A. (D) Levels of SUMOylated MAGI1 were quantified by densitometry of immunoblots from 3 independent experiments and are plotted (n = 3). Data represent mean ± SEM. **P < 0.01. (E) The levels of SUMOylated MAGI1, MAGI1, and hemagglutinin-tagged (HA-tagged) SUMO3 expression in ECs coexpressing HA-tagged pcDNA-SUMO3 and pCMV-Flag-MAGI1-WT or -K931R determined as in A. (F and G) HUVECs were transduced with Ad-Flag-MAGI1 WT or Ad-Flag-MAGI1 K931R mutant with His-tagged SUMO3 or a control vector. His-tagged SUMO3 was pulled down by Dynabeads and immunoblotted with anti-Flag (MAGI1) and anti-His (SUMO3). (G) Quantification of pulled down SUMOylated MAGI1 expression relative to that of Flag-MAGI1 (n = 3). Data represent mean ± SEM. **P < 0.01. (H) ECs expressing pCMV-Flag-MAGI1-WT or -K931R or a control vector were subjected to an NF-κB activity assay with TNF-α treatment (10 ng/ml, 24 h). The relative NF-κB luciferase activity was measured and presented as the mean ± SEM (n = 6). **P < 0.01. (I) Bovine aortic ECs expressing pCMV-Flag–tagged MAGI1-WT or -K931R were exposed to d-flow for 12 hours, and the levels of E selectin, ICAM-1, and MAGI1 expression were determined. (J) The intensity of ICAM-1 expression relative to that of tubulin expression was quantified. Data represent mean ± SEM (n = 3). **P < 0.01; ***P < 0.001. Statistical differences between 2 independent groups (G) were assessed using the Student’s t test (2-tailed) and 1-way ANOVA followed by Bonferroni’s post hoc testing for multiple groups (B, D, H, and J).