ABSTRACT
Nonvesicular transport of cholesterol plays an essential role in the distribution and regulation of cholesterol within cells, but it has been difficult to identify the key intracellular cholesterol transporters. The steroidogenic acute regulatory-related lipid-transfer (START) family of proteins is involved in several pathways of nonvesicular trafficking of sterols. Among them, STARD4 has been shown to increase intracellular cholesteryl ester formation and is controlled at the transcriptional level by sterol levels in cells. We found that STARD4 is very efficient in transporting sterol between membranes in vitro. Cholesterol levels are increased in STARD4-silenced cells, while sterol transport to the endocytic recycling compartment (ERC) and to the endoplasmic reticulum (ER) are enhanced upon STARD4 overexpression. STARD4 silencing attenuates cholesterol-mediated regulation of SREBP-2 activation, while its overexpression amplifies sterol sensing by SCAP/SREBP-2. To analyze STARD4's mode of action, we compared sterol transport mediated by STARD4 with that of a simple sterol carrier, methyl-ß-cyclodextrin (MCD), when STARD4 and MCD were overexpressed or injected into cells. Interestingly, STARD4 and cytosolic MCD act similarly by increasing the rate of transfer of sterol to the ERC and to the ER. Our results suggest that cholesterol transport mediated by STARD4 is an important component of the cholesterol homeostasis regulatory machinery.
Subject(s)
Cholesterol/metabolism , Membrane Transport Proteins/metabolism , Amino Acid Motifs , Cell Line, Tumor , Cell Membrane/metabolism , Cholesterol Esters/biosynthesis , Endoplasmic Reticulum/metabolism , Ergosterol/analogs & derivatives , Ergosterol/metabolism , Esterification , Fluorescence Recovery After Photobleaching , Fluorescent Dyes/metabolism , Gene Knockdown Techniques , Homeostasis , Humans , Intracellular Signaling Peptides and Proteins/metabolism , Kinetics , Liposomes/metabolism , Membrane Proteins/metabolism , Membrane Transport Proteins/chemistry , Membrane Transport Proteins/genetics , Protein Structure, Tertiary , RNA Interference , Sterol O-Acyltransferase/antagonists & inhibitors , Sterol O-Acyltransferase/metabolism , Sterol Regulatory Element Binding Protein 2/metabolism , Time-Lapse Imaging , Transferrin/metabolism , Transport Vesicles/metabolism , beta-Cyclodextrins/pharmacologyABSTRACT
Recent studies have shown that cooperative interactions in endoplasmic reticulum (ER) membranes between Scap, cholesterol, and Insig result in switch-like control over activation of SREBP-2 transcription factors. This allows cells to rapidly adjust rates of cholesterol synthesis and uptake in response to even slight deviations from physiological set-point levels, thereby ensuring cholesterol homeostasis. In the present study we directly probe for the accessibility of cholesterol in purified ER membranes. Using a soluble cholesterol-binding bacterial toxin, perfringolysin O, we show that cholesterol accessibility increases abruptly at â¼5 mol % ER cholesterol, the same concentration at which SREBP-2 activation is halted. This switch-like change in cholesterol accessibility is observed not only in purified ER membranes but also in liposomes made from ER lipid extracts. The accessibility of cholesterol in membranes is related to its chemical activity. Complex formation between cholesterol and some ER phospholipids can result in sharp changes in cholesterol chemical activity and its accessibility to perfringolysin O or membrane sensors like Scap. The control of the availability of the cholesterol ligand to participate in cooperative Scap/cholesterol/Insig interactions further sharpens the sensitive switch that exerts precise control over cholesterol levels in cell membranes.
Subject(s)
Cholesterol/chemistry , Endoplasmic Reticulum/chemistry , Sterol Regulatory Element Binding Protein 2/metabolism , Animals , Bacterial Toxins/chemistry , Bacterial Toxins/metabolism , CHO Cells , Cricetinae , Cricetulus , Endoplasmic Reticulum/metabolism , Hemolysin Proteins/chemistry , Hemolysin Proteins/metabolism , Immunoblotting , Liposomes/chemistry , Phospholipids/chemistry , Protein Binding , Sterol Regulatory Element Binding Protein 2/chemistryABSTRACT
Estrogen hormones play critical roles in the regulation of many tissue functions. The effects of estrogens are primarily mediated by the estrogen receptors (ER) alpha and beta. ERs are ligand-activated transcription factors that regulate a complex array of genomic events that orchestrate cellular growth, differentiation and death. Although many factors contribute to their etiology, estrogens are thought to be the primary agents for the development and/or progression of target tissue malignancies. Many of the current modalities for the treatment of estrogen target tissue malignancies are based on agents with diverse pharmacology that alter or prevent ER functions by acting as estrogen competitors. Although these compounds have been successfully used in clinical settings, the efficacy of treatment shows variability. An increasing body of evidence implicates ERalpha polymorphisms as one of the contributory factors for differential responses to estrogen competitors. This review aims to highlight the recent findings on polymorphisms of the lately identified ERbeta in order to provide a functional perspective with potential pharmacogenomic implications.
