Cholesterol Binding at the Cholesterol Recognition/Interaction Amino Acid Consensus (CRAC) of the Peripheral-Type Benzodiazepine Receptor and Inhibition of Steroidogenesis by an HIV TAT-CRAC Peptide
We previously defined a cholesterol recognition/interaction amino acid consensus (CRAC; ATVLNYYVWRDNS in the carboxyl terminus of the peripheral-type benzodiazepine receptor (PBR), an outer mitochondrial membrane protein involved in the regulation of cholesterol transport into the mitochondria, the...
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Published in | Proceedings of the National Academy of Sciences - PNAS Vol. 98; no. 3; pp. 1267 - 1272 |
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Main Authors | , , , , |
Format | Journal Article |
Language | English |
Published |
United States
National Academy of Sciences
30.01.2001
National Acad Sciences The National Academy of Sciences |
Subjects | |
Online Access | Get full text |
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Summary: | We previously defined a cholesterol recognition/interaction amino acid consensus (CRAC; ATVLNYYVWRDNS in the carboxyl terminus of the peripheral-type benzodiazepine receptor (PBR), an outer mitochondrial membrane protein involved in the regulation of cholesterol transport into the mitochondria, the rate-determining step in steroid biosynthesis. We examined (i) the PBR-cholesterol interaction by UV crosslinking of the C17 side-chain containing progestin, promegestone, and (ii) the role of the CRAC domain of PBR in Leydig cell steroidogenesis by using a transducible peptide composed of the TAT domain of HIV and the CRAC domain of PBR. [3H]Promegestone photoincorporated into recombinant PBR, and this labeling was displaced by cholesterol. [3H]Promegestone also photoincorporated into the TAT-CRAC peptide. [3H]Promegestone crosslinking to TAT-CRAC could be displaced by cholesterol and promegestone, with IC50 values of 1 and 200 µM, respectively. TAT-CRAC efficiently transduced into MA-10 Leydig cells and inhibited the hCG- and cAMP-stimulated steroid production in a dose-dependent manner. TAT-CRAC did not affect the hCG-induced cAMP synthesis and the 22R-hydroxycholesterol-supported steroidogenesis. Mutated TAT-CRAC lost its ability to bind [3H]promegestone and to inhibit the hCG-stimulated steroidogenesis. These results show that TAT-CRAC binds cholesterol and competes for cholesterol interaction with endogenous PBR, suggesting that the cytosolic carboxyl-terminal domain of PBR is responsible for taking up and bringing steroidogenic cholesterol into the mitochondria. |
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Bibliography: | ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 Edited by Erminio Costa, University of Illinois, Chicago, IL, and approved December 6, 2000 On leave from the Institute of Pharmaceutical Biology, Martin-Luther-University Halle-Wittenberg, Halle/Saale, Germany. To whom reprint requests should be addressed. E-mail: papadopv@gunet.georgetown.edu. |
ISSN: | 0027-8424 1091-6490 |
DOI: | 10.1073/pnas.031461598 |