Macrophage SR-BI modulates autophagy via VPS34 complex and PPARα transcription of Tfeb in atherosclerosis
Huan Tao, Patricia G. Yancey, John L. Blakemore, Youmin Zhang, Lei Ding, W. Gray Jerome, Jonathan D. Brown, Kasey C. Vickers, MacRae F. Linton
Huan Tao, Patricia G. Yancey, John L. Blakemore, Youmin Zhang, Lei Ding, W. Gray Jerome, Jonathan D. Brown, Kasey C. Vickers, MacRae F. Linton
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Research Article Cardiology Vascular biology

Macrophage SR-BI modulates autophagy via VPS34 complex and PPARα transcription of Tfeb in atherosclerosis

Abstract

Autophagy modulates lipid turnover, cell survival, inflammation, and atherogenesis. Scavenger receptor class B type I (SR-BI) plays a crucial role in lysosome function. Here, we demonstrate that SR-BI regulates autophagy in atherosclerosis. SR-BI deletion attenuated lipid-induced expression of autophagy mediators in macrophages and atherosclerotic aortas. Consequently, SR-BI deletion resulted in 1.8- and 2.5-fold increases in foam cell formation and apoptosis, respectively, and increased oxidized LDL–induced inflammatory cytokine expression. Pharmacological activation of autophagy failed to reduce lipid content or apoptosis in Sr-b1–/– macrophages. SR-BI deletion reduced both basal and inducible levels of transcription factor EB (TFEB), a master regulator of autophagy, causing decreased expression of autophagy genes encoding VPS34 and Beclin-1. Notably, SR-BI regulated Tfeb expression by enhancing PPARα activation. Moreover, intracellular macrophage SR-BI localized to autophagosomes, where it formed cholesterol domains resulting in enhanced association of Barkor and recruitment of the VPS34–Beclin-1 complex. Thus, SR-BI deficiency led to lower VPS34 activity in macrophages and in atherosclerotic aortic tissues. Overexpression of Tfeb or Vps34 rescued the defective autophagy in Sr-b1–/– macrophages. Taken together, our results show that macrophage SR-BI regulates autophagy via Tfeb expression and recruitment of the VPS34–Beclin-1 complex, thus identifying previously unrecognized roles for SR-BI and potentially novel targets for the treatment of atherosclerosis.

Authors

Huan Tao, Patricia G. Yancey, John L. Blakemore, Youmin Zhang, Lei Ding, W. Gray Jerome, Jonathan D. Brown, Kasey C. Vickers, MacRae F. Linton

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Figure 11

SR-BI deficiency decreases PPARα activity in macrophages.

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SR-BI deficiency decreases PPARα activity in macrophages. (A) PPARα acti...
(A) PPARα activity was measured using serial doses of nuclear extracts (NEs) from WT and Sr-b1–/– macrophages treated with 100 μg/mL acetylated LDL and 5 μg/mL Sandoz 58035 (FC enriched) for 24 hours. The data are expressed as mean ± SEM from 3 independent experiments (n = 3 per group). *P < 0.05, **P < 0.01, ***P < 0.001 by unpaired Student’s t test. (B) WT and Sr-b1–/– cells were FC enriched and treated with or without either CP-775146 or GW6471. The levels of PPARα, TFEB, and c-Jun in nuclear extracts were then detected by Western blotting. The blots are representative, and the numbers are the mean of 2 experiments, in which the values are normalized to either basal WT (green, regular font) or basal Sr-b1–/– levels (red, italic font). (C) WT and Sr-b1–/– macrophages treated with 10% FBS or enriched with FC by incubation with acetylated LDL and Sandoz 58035 for 24 hours. The expression of mRNA encoding PPARα was then measured by real-time PCR. (D) The expression levels of mRNA for TFEB, Beclin-1, Rab7, LC3, VPS34, and ATG5 were determined by real-time PCR in FC-enriched WT and Sr-b1–/– macrophages treated with vehicle alone or with either CP-775146 or GW6471. (E) The relative PPARα mRNA levels were measured by real-time PCR in FC-enriched WT and Sr-b1–/– macrophages treated with vehicle alone or with CP-775146 or GW6471. In C–E, the data are expressed as mean ± SEM from 3 independent experiments (n = 3 per group). *P < 0.05, **P < 0.01, ***P < 0.001 by 1-way ANOVA with Bonferroni’s post hoc test. NS, not significant. (F) The expression of Beclin-1, TFEB, VPS34, LC3I, LC3II, p62, Rab7, and GAPDH was analyzed by Western blotting in FC-enriched WT and Sr-b1–/– macrophages treated with vehicle, CP-775146, or GW6471. The blots are representative, and the numbers are the mean of 2 independent experiments. In B and F, the values are normalized to either basal WT (green, regular font) or basal Sr-b1–/– levels (red, italic font).