An intracellular role for ABCG1-mediated cholesterol transport in the regulated secretory pathway of mouse pancreatic β cells
Jeffrey M. Sturek, … , Raghavendra G. Mirmira, Catherine C. Hedrick
Jeffrey M. Sturek, … , Raghavendra G. Mirmira, Catherine C. Hedrick
Published June 7, 2010
Citation Information: J Clin Invest. 2010;120(7):2575-2589. https://doi.org/10.1172/JCI41280.
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Research Article Metabolism

An intracellular role for ABCG1-mediated cholesterol transport in the regulated secretory pathway of mouse pancreatic β cells

Abstract

Cholesterol is a critical component of cell membranes, and cellular cholesterol levels and distribution are tightly regulated in mammals. Recent evidence has revealed a critical role for pancreatic β cell–specific cholesterol homeostasis in insulin secretion as well as in β cell dysfunction in diabetes and the metabolic response to thiazolidinediones (TZDs), which are antidiabetic drugs. The ATP-binding cassette transporter G1 (ABCG1) has been shown to play a role in cholesterol efflux, but its role in β cells is currently unknown. In other cell types, ABCG1 expression is downregulated in diabetes and upregulated by TZDs. Here we have demonstrated an intracellular role for ABCG1 in β cells. Loss of ABCG1 expression impaired insulin secretion both in vivo and in vitro, but it had no effect on cellular cholesterol content or efflux. Subcellular localization studies showed the bulk of ABCG1 protein to be present in insulin granules. Loss of ABCG1 led to altered granule morphology and reduced granule cholesterol levels. Administration of exogenous cholesterol restored granule morphology and cholesterol content and rescued insulin secretion in ABCG1-deficient islets. These findings suggest that ABCG1 acts primarily to regulate subcellular cholesterol distribution in mouse β cells. Furthermore, islet ABCG1 expression was reduced in diabetic mice and restored by TZDs, implicating a role for regulation of islet ABCG1 expression in diabetes pathogenesis and treatment.

Authors

Jeffrey M. Sturek, J. David Castle, Anthony P. Trace, Laura C. Page, Anna M. Castle, Carmella Evans-Molina, John S. Parks, Raghavendra G. Mirmira, Catherine C. Hedrick

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Figure 3

ABCG1 deficiency does not alter total β cell cholesterol content or efflux.

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ABCG1 deficiency does not alter total β cell cholesterol content or effl...
(A) MIN6 cells were labeled with [3H]cholesterol overnight, and [3H]cholesterol efflux to extracellular acceptors over 4 hours was measured (n = 6). (B) Western blotting showing siRNA-mediated knockdown of ABCG1 in MIN6 cells (approximately 90% by densitometry). (C, D) MIN6 cells were treated with siRNA as in B and a time-course (C) and dose-response curve (D) of HDL-mediated [3H]cholesterol efflux was measured (n = 4; representative of 3 independent experiments). (E) ApoAI-mediated cholesterol efflux was measured in MIN6 cells treated with ABCG1 siRNA. (F) Islets were isolated from WT and Abcg1–/– mice and total cholesterol (TC), free-cholesterol (FC), and cholesterol-ester (CE) content were measured by gas chromatography (150 islets per sample, n = 14 mice per group). (G) qRT-PCR was performed on isolated islets for cholesterol- (SREBP-2, LDLR, HMG-CoA R, HMG-CoA S) and oxysterol-regulated (ABCA1, SREBP-1c) mRNA (n = 5–10 mice per genotype). (H) Triglyceride content was measured in isolated islets from WT and Abcg1–/– mice (200 islets per sample, n = 8 mice per genotype). LDLR, LDL receptor; HMG-CoA R, HMG-CoA reductase; HMG-CoA S, HMG-CoA synthase. Data are presented as mean ± SEM. ***P < 0.001.