Cholesterol transport in steroid biosynthesis: role of protein-protein interactions and implications in disease states

Abstract

The transfer of cholesterol from the outer to the inner mitochondrial membrane is the rate-limiting step in hormone-induced steroid formation. To ensure that this step is achieved efficiently, free cholesterol must accumulate in excess at the outer mitochondrial membrane and then be transferred to the inner membrane. This is accomplished through a series of steps that involve various intracellular organelles, including lysosomes and lipid droplets, and proteins such as the translocator protein (18 kDa, TSPO) and steroidogenic acute regulatory (StAR) proteins. TSPO, previously known as the peripheral-type benzodiazepine receptor, is a high-affinity drug- and cholesterol-binding mitochondrial protein. StAR is a hormone-induced mitochondria-targeted protein that has been shown to initiate cholesterol transfer into mitochondria. Through the assistance of proteins such as the cAMP-dependent protein kinase regulatory subunit Ialpha (PKA-RIalpha) and the PKA-RIalpha- and TSPO-associated acyl-coenzyme A binding domain containing 3 (ACBD3) protein, PAP7, cholesterol is transferred to and docked at the outer mitochondrial membrane. The TSPO-dependent import of StAR into mitochondria, and the association of TSPO with the outer/inner mitochondrial membrane contact sites, drives the intramitochondrial cholesterol transfer and subsequent steroid formation. The focus of this review is on (i) the intracellular pathways and protein-protein interactions involved in cholesterol transport and steroid biosynthesis and (ii) the roles and interactions of these proteins in endocrine pathologies and neurological diseases where steroid synthesis plays a critical role.

Figure 1. Trafficking of cholesterol to the mitochondria for steroidogenesis

Pathway 1: Cholesterol synthesized in the ER is trafficked to the Golgi apparatus where it can be targeted to the mitochondria via the PAP7 protein for steroidogenesis. Passive diffusion from the ER to the mitochondria, shown here with the corresponding cholesterol molecules present on both organelles, is another possible pathway for steroidogenic cholesterol to be transferred to the mitochondria. Pathway 2: Low density lipoprotein (LDL), containing cholesterol, binds to the LDL receptor and is trafficked through the endosomal pathway. MLN-64 assists with the transfer of cholesterol to the mitochondria from the late endosomes and lysosomes for use in steroidogenesis. NPC1 and NPC2 associate with MLN-64 for cholesterol transfer out of the lysosomes, although it is not known if this cholesterol is used for steroidogenesis. Pathway 3: Cholesterol is transferred from high density lipoprotein (HDL) to the plasma membrane by the SR-BI receptor. Hormone-sensitive lipase (HSL) converts esterified cholesterol from the plasma membrane to free cholesterol, which can be used for steroidogenesis. Pathway 4: HSL also interacts with esterified cholesterol present in the lipid droplets (LD), which converts esterified cholesterol to free cholesterol for use in steroidogenesis as well. Free cholesterol from the LD can interact with lipid-binding proteins present in the cytosol for delivery to the mitochondria.

Figure 2. Cholesterol-binding domains of StAR and TSPO

A) Human StAR protein modeled with cholesterol (brown) present in the sterol-binding domain (PDB ID code: 2I93). Homology models are deduced via the Swiss-model services under project (optimize) mode using the crystal structures of human MLN64 (PDB ID code: 1EM2) as template (http://swissmodel.expasv.org/SWISS-MODEL.html. B) Molecular model of TSPO’s five alpha helices in the presence of cholesterol, demonstrating a pore accommodating a cholesterol molecule. Homology modeling of mouse TSPO was performed using apolipophorin-III from Manduca sexta as template (PDB ID code: 1EQ1) [158]. C) Ribbon diagram of mTSPO, showing the five helices as well as the CRAC domain, consisting of Leu/Tyr/Arg residues, and cholesterol (top view). D) Docking model of cholesterol to the TSPO CRAC domain. The accessible surface of the peptide and cholesterol molecules are represented in red and blue, respectively (Reprinted with permission from [159]).

Figure 3. Protein-protein interactions at the OMM

A) Proposed basal-state protein-protein interactions of TSPO, VDAC, and ANT. B) Transduceosome complex formed after acute hormonal stimulation. The TSPO, PAP7 (ACBD3), and VDAC complex recruits PKA-RIα, which is shown binding to PAP7 (ACBD3). The accumulation of cAMP activates PKA-RIα, releasing the catalytic subunits and phosphorylating StAR present at the OMM. StAR interacts with both TSPO and VDAC and is imported into the IMM. Cholesterol is imported into the IMM with the assistance of the transduceosome complex and the presence of DBI; there the cholesterol interacts with the CYP11A1 side chain cleavage enzyme to be converted to pregnenolone. ANT is represented in the IMM as identified from previous isolated complexes with TSPO.