Role of protein-protein interactions in mitochondrial protein import, cholesterol transport and steroid biosynthesis

Steroid synthesis is initiated by the transfer of cholesterol to the inner membrane of the mitochondria where the conversion of cholesterol to pregnenolone occurs through the C27 cholesterol side chain cleavage cytochrome P450 enzyme (P450scc CYP11A1). The rate of steroidogenesis is not regulated by the activity of CYP11A1 but by the availability of substrate. As the process of trafficking cholesterol to the mitochondria can occur through a series of cytosolic and mitochondrial proteins and vesicular interactions, we propose that this delivery of cholesterol into the mitochondria occurs through specific protein-protein interactions that drive steroidogenesis.This hypothesis was explored using MA-10 mouse Leydig cells, which undergo steroidogenesis to produce progesterone. It was identified through cross-linking studies that a protein complex formed at the outer mitochondrial membrane (OMM) upon hormonal stimulation. This complex consisted of mitochondrial protein translocator protein (TSPO, 18 kDa), Golgi protein PBR Associated Protein 7, (PAP7), cytosolic protein PKA-RIalpha, and mitochondrially targeted Steroidogenic Acute Regulatory protein (StAR). TSPO assists with the translocation of cholesterol from the OMM to the IMM and is an integral OMM protein therefore, we decided to study the mechanisms of its import and integration into the OMM. We identified that the C-terminus and amino acids In summary, these studies demonstrate that delivery of cholesterol into the mitochondria and thus steroidogenesis are driven by a series of protein-protein interactions.