Aster proteins regulate the accessible cholesterol pool in the plasma membrane.

Ferrari, Alessandra; He, Cuiwen; Kennelly, John Paul; Sandhu, Jaspreet; Xiao, Xu; Chi, Xun; Jiang, Haibo; Young, Stephen G; Tontonoz, Peter

Recent studies have demonstrated the existence of a discrete pool of cholesterol in the plasma membranes (PM) of mammalian cells--referred to as the accessible cholesterol pool-that can be detected by the binding of modified versions of bacterial cytolysins (e.g, anthrolysin O). When the amount of accessible cholesterol in the PM exceeds a threshold level, the excess cholesterol moves to the ER where it regulates the SREBP2 pathway and undergoes esterification. We reported previously that the Aster/Gramd1 family of sterol transporters mediates nonvesicular movement of cholesterol from the PM to endoplasmic reticulum (ER) in multiple mammalian cell types. Here, we investigated the PM pool of accessible cholesterol in cholesterol-loaded fibroblasts with a knockdown of Aster-A and in mouse macrophages from Aster-B and Aster-A/B-deficient mice. Nanoscale secondary ion mass spectrometry (NanoSIMS) analyses revealed an expansion of the accessible cholesterol pool in cells lacking Aster expression. The increased accessible cholesterol pool in the PM was accompanied by reduced cholesterol to the ER, evident by increased expression of SREBP2-regulated genes. Co-sedimentation experiments with liposomes revealed that the Aster-B GRAM domain binds to membranes in a cholesterol concentration-dependent manner and that the binding is facilitated by the presence of phosphatidylserine. These studies reveal that the Aster-mediated nonvesicular cholesterol transport pathway controls levels of accessible cholesterol in the PM as well as the activity of the SREBP pathway.