Abstract
When human fibroblasts take up plasma low density lipoprotein (LDL), its cholesterol is liberated in lysosomes and eventually reaches the endoplasmic reticulum (ER) where it inhibits cholesterol synthesis by blocking activation of SREBPs. This feedback protects against cholesterol overaccumulation in the plasma membrane (PM). But how does ER know whether PM is saturated with cholesterol? In this study, we define three pools of PM cholesterol: (1) a pool accessible to bind 125I-PFO*, a mutant form of bacterial Perfringolysin O, which binds cholesterol in membranes; (2) a sphingomyelin(SM)-sequestered pool that binds 125I-PFO* only after SM is destroyed by sphingomyelinase; and (3) a residual pool that does not bind 125I-PFO* even after sphingomyelinase treatment. When LDL-derived cholesterol leaves lysosomes, it expands PM's PFO-accessible pool and, after a short lag, it also increases the ER's PFO-accessible regulatory pool. This regulatory mechanism allows cells to ensure optimal cholesterol levels in PM while avoiding cholesterol overaccumulation.DOI: http://dx.doi.org/10.7554/eLife.02882.001
Highlights
Animal cells tightly control the level of cholesterol in their plasma membranes (PMs)
When endoplasmic reticulum (ER) cholesterol is less than ∼5 mole% of total ER lipids (Radhakrishnan et al, 2008), the Scap/SREBP-2 complex enters COPII-coated vesicles that move to the Golgi, where two proteases liberate the active fragment of SREBP-2 (Sun et al, 2007)
These data raise the possibility that low density lipoprotein (LDL)-derived cholesterol must first expand a pool of cholesterol in the PM before it is delivered to the ER in sufficient amounts to undergo esterification by acyl-CoA:cholesterol acyltransferase (ACAT)
Summary
Animal cells tightly control the level of cholesterol in their plasma membranes (PMs). When ER cholesterol is less than ∼5 mole% of total ER lipids (Radhakrishnan et al, 2008), the Scap/SREBP-2 complex enters COPII-coated vesicles that move to the Golgi, where two proteases liberate the active fragment of SREBP-2 (Sun et al, 2007). The active fragment enters the nucleus where it activates transcription of the cholesterolsynthesizing genes and the gene for the low density lipoprotein (LDL) receptor, which supplies the cell with exogenous cholesterol (Brown and Goldstein, 1997). When the ER cholesterol rises above a sharp threshold of 5 mole% of total ER lipids (Radhakrishnan et al, 2008), the Scap/SREBP complex binds to an ER anchor protein called Insig, and this prevents its transport to the Golgi (Goldstein et al, 2006). It is removed from the PM and delivered to the ER where it is esterified by acyl-CoA:cholesterol acyltransferase (ACAT) for storage as cytoplasmic cholesteryl ester droplets (Brown and Goldstein, 1986)
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