Disruption of USP9X in macrophages promotes foam cell formation and atherosclerosis
Biqing Wang, … , Hongfeng Jiang, Ding Ai
Biqing Wang, … , Hongfeng Jiang, Ding Ai
Published April 7, 2022
Citation Information: J Clin Invest. 2022;132(10):e154217. https://doi.org/10.1172/JCI154217.
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Disruption of USP9X in macrophages promotes foam cell formation and atherosclerosis

Abstract

Subendothelial macrophage internalization of modified lipids and foam cell formation are hallmarks of atherosclerosis. Deubiquitinating enzymes (DUBs) are involved in various cellular activities; however, their role in foam cell formation is not fully understood. Here, using a loss-of-function lipid accumulation screening, we identified ubiquitin-specific peptidase 9 X-linked (USP9X) as a factor that suppressed lipid uptake in macrophages. We found that USP9X expression in lesional macrophages was reduced during atherosclerosis development in both humans and rodents. Atherosclerotic lesions from macrophage USP9X-deficient mice showed increased macrophage infiltration, lipid deposition, and necrotic core content than control apolipoprotein E–KO (Apoe–/–) mice. Additionally, loss-of-function USP9X exacerbated lipid uptake, foam cell formation, and inflammatory responses in macrophages. Mechanistically, the class A1 scavenger receptor (SR-A1) was identified as a USP9X substrate that removed the K63 polyubiquitin chain at the K27 site. Genetic or pharmacological inhibition of USP9X increased SR-A1 cell surface internalization after binding of oxidized LDL (ox-LDL). The K27R mutation of SR-A1 dramatically attenuated basal and USP9X knockdown–induced ox-LDL uptake. Moreover, blocking binding of USP9X to SR-A1 with a cell-penetrating peptide exacerbated foam cell formation and atherosclerosis. In this study, we identified macrophage USP9X as a beneficial regulator of atherosclerosis and revealed the specific mechanisms for the development of potential therapeutic strategies for atherosclerosis.

Authors

Biqing Wang, Xuening Tang, Liu Yao, Yuxin Wang, Zhipeng Chen, Mengqi Li, Naishi Wu, Dawei Wu, Xiangchen Dai, Hongfeng Jiang, Ding Ai

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Figure 8

An inhibitory peptide blocks the SR-A1–USP9X interaction and promotes foam cell formation and atherosclerosis.

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An inhibitory peptide blocks the SR-A1–USP9X interaction and promotes fo...
(A) HEK293 cells were cotransfected with constructs expressing the middle region of USP9X (FLAG-USP9X-M) and MYC tagged different truncations of the intracellular segment of SR-A1 before IP of whole cell lysates with FLAG magnetic beads (n = 5). (B) BMDMs were treated with different peptides (20 μM) for 24 hours (n = 5). The cells were subjected to IP with antibody against USP9X or IgG to quantify the interaction of USP9X and SR-A1. (C) Oil Red O staining analysis was performed. (D) Oil Red O staining of aortas from control or inhibitory peptides administered to Apoe–/– mice fed WD for 10 weeks (n = 10). Scale bar: 5 mm. (E) Quantification of plaque area/total vessel area. AA, aortic arch; TA, thoracic aorta. Unpaired 2-tailed Student’s t test (n = 10). (F) Representative images of H&E staining of aortic root sections (left) and quantification of lesion and necrotic core areas (right). Black dashed lines demarcate atherosclerotic plaques. Scale bar: 100 μm. Unpaired 2-tailed Student’s t test (n = 10). (G) Oil Red O (left) staining of aortic root sections. Scale bar: 100 μm. Macrophages identified by anti-CD68 antibody staining (right). The white dashed line indicates plaques. Scale bar: 50 μm. Unpaired 2-tailed Student’s t test (n = 10).