title: Cholesterol accumulation inhibits ER to Golgi transport and protein secretion: studies of apolipoprotein E and VSVGt.
abstract: Cholesterol excess is typical of various diseases including atherosclerosis. We have investigated whether cholesterol accumulation in the ER (endoplasmic reticulum) can inhibit exit of vesicular cargo and secretion of proteins by studying apoE (apolipoprotein E), a significant glycoprotein in human health and disease. CHO (Chinese hamster ovary) cells expressing human apoE under a cholesterol-independent promoter incubated with cholesterol-cyclodextrin complexes showed increased levels of cellular free and esterified cholesterol, inhibition of SREBP-2 (sterol-regulatory-element-binding protein 2) processing, and a mild induction of ER stress, indicating significant accumulation of cholesterol in the ER. Secretion of apoE was markedly inhibited by cholesterol accumulation, and similar effects were observed in cells enriched with lipoprotein-derived cholesterol and in primary human macrophages. Removal of excess cholesterol by a cyclodextrin vehicle restored apoE secretion, indicating that the transport defect was reversible. That cholesterol impaired protein trafficking was supported by the cellular accumulation of less sialylated apoE glycoforms, and by direct visualization of altered ER to Golgi transport of thermo-reversible VSVG (vesicular stomatitis virus glycoprotein) linked to GFP (green fluorescent protein). We conclude that intracellular accumulation of cholesterol in the ER reversibly inhibits protein transport and secretion. Strategies to correct ER cholesterol may restore homoeostatic processes and intracellular protein transport in conditions characterized by cholesterol excess.
authors: Centre for Vascular Research, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia.
authors: Maaike
authors: M
authors: Kockx
authors: Donna L
authors: DL
authors: Dinnes
authors: Kuan-Yen
authors: KY
authors: Huang
authors: Laura J
authors: LJ
authors: Sharpe
authors: Wendy
authors: W
authors: Jessup
authors: Andrew J
authors: AJ
authors: Brown
authors: Leonard
authors: L
authors: Kritharides
mesh_terms: D000818:Animals; D001057:Apolipoproteins E / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D002784:Cholesterol / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D059865:Endoplasmic Reticulum Stress; D006031:Glycosylation; D006056:Golgi Apparatus / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012794:Sialic Acids / Q000378:metabolism; D018116:Vesiculovirus / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000378:metabolism*
chemical_list: D001057:Apolipoproteins E; C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D012794:Sialic Acids; D014759:Viral Envelope Proteins; D049452:Green Fluorescent Proteins; D002784:Cholesterol