Peripheral-type benzodiazepine receptor function in cholesterol transport. Identification of a putative cholesterol recognition/interaction amino acid sequence and consensus pattern.
In steroid-synthesizing cells, like the MA-10 mouse tumor Leydig cells, the peripheral-type benzodiazepine receptor (PBR) is an outer mitochondrial membrane protein involved in the regulation of cholesterol transport from the outer to the inner mitochondrial membrane, the rate-determining step in steroid biosynthesis. Expression of PBR in Escherichia coli DE3 cells, which have no PBR, no cholesterol, and do not make steroids, induced the ability to take up cholesterol in a time-dependent, temperature-sensitive, and energy-independent manner. These cells took up no other steroids tested. Addition of the high affinity PBR ligand PK 11195 to cholesterol-loaded membranes, obtained from cells transfected with PBR, resulted in the release of the uptaken cholesterol. Expression in DE3 cells of mutant PBRs demonstrated that deletions in the cytoplasmic carboxy-terminus dramatically reduced the cholesterol uptake function of PBR, although it retained full capacity to bind PK 11195. Site-directed mutagenesis in the carboxy-terminal region of PBR demonstrated that bacteria expressing the mutant PBR proteins PBR(Y153S) and PBR(R156L) do not accumulate cholesterol, suggesting that amino acids Y153 and R156 are involved in the interaction of the receptor with cholesterol. Considering these results, we postulate the existence of a common cholesterol recognition/interaction amino acid consensus pattern (-L/V-(X)(1-5)-Y-(X)(1-5)-R/K-). Indeed, we found this amino acid consensus pattern in all proteins shown to interact with cholesterol. In conclusion, these data suggest that the expression of PBR confers the ability to take up and release, upon ligand activation, cholesterol. Considering the widespread occurrence of this protein and its tissue and cell specific subcellular localization, these results suggest a more general role of PBR in intracellular cholesterol transport and compartmentalization.
Li
H
H
Department of Cell Biology, Georgetown University Medical Center, Washington, DC 20007, USA.
Papadopoulos
V
V
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002784:Cholesterol / Q000378:metabolism*; D016384:Consensus Sequence; D004926:Escherichia coli / Q000378:metabolism; D017353:Gene Deletion; D007546:Isoquinolines / Q000378:metabolism; D008024:Ligands; D051379:Mice; D009154:Mutation / Q000502:physiology; D011963:Receptors, GABA-A / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured
D007546:Isoquinolines; D008024:Ligands; D011963:Receptors, GABA-A; D011994:Recombinant Proteins; D002784:Cholesterol; C037850:PK 11195