Adrenal Mitochondria and Steroidogenesis: From Individual Proteins to Functional Protein Assemblies

Abstract

The adrenal cortex is critical for physiological function as the central site of glucocorticoid and mineralocorticoid synthesis. It possesses a great degree of specialized compartmentalization at multiple hierarchical levels, ranging from the tissue down to the molecular levels. In this paper, we discuss this functionalization, beginning with the tissue zonation of the adrenal cortex and how this impacts steroidogenic output. We then discuss the cellular biology of steroidogenesis, placing special emphasis on the mitochondria. Mitochondria are classically known as the "powerhouses of the cell" for their central role in respiratory adenosine triphosphate synthesis, and attention is given to mitochondrial electron transport, in both the context of mitochondrial respiration and mitochondrial steroid metabolism. Building on work demonstrating functional assembly of large protein complexes in respiration, we further review research demonstrating a role for multimeric protein complexes in mitochondrial cholesterol transport, steroidogenesis, and mitochondria-endoplasmic reticulum contact. We aim to highlight with this review the shift in steroidogenic cell biology from a focus on the actions of individual proteins in isolation to the actions of protein assemblies working together to execute cellular functions.

Keywords: cholesterol transport; cytochrome P450 enzyme system; endoplasmic reticulum; mitochondria; steroidogenic acute regulatory protein; translocator protein; voltage-dependent anion channels.

Figure 1

Schematic of adrenal zonation. The different functional zones of the human adrenal gland are depicted with the outermost capsule layer overlying the mineralocorticoid-synthesizing glomerulosa layer. The fasciculata layer lies under the granulosa layer and is responsible for the synthesis of the glucocorticoid cortisol. The final layer of the cortex, the reticularis, synthesizes the androgen dehydroepiandrosterone (DHEA), while the innermost layer of the schematic, the medulla, is composed of chromaffin cells, responsible for the production of the catecholamine epinephrine.

Figure 2

Schematics of adrenal steroidogenic pathways. The metabolism of cholesterol to pregnenolone by the mitochondrial CYP11A1 is common to all three zones of the human adrenal. (A) The mitochondrial/microsomal enzyme HSD3B converts pregnenolone to progesterone, which is metabolized to 11-deoxycorticosterone by the microsomal CYP21. The final reactions of aldosterone synthesis are catalyzed by the mitochondrial CYP11B2, which converts 11-deoxycorticosterone to corticosterone, which is hydroxylated at C18 to form 18-hydroxycorticosterone which is then finally converted to aldosterone. (B) In the zona fasciculata, the microsomal CYP17 and the mitochondrial/microsomal HSD3B can generate 17-hydroxyprogesterone, progesterone, and 17-hydroxyprogesterone. The microsomal CYP21 preferentially metabolizes 17-hydroxyprogesterone to 11-deoxycortisol, which is finally metabolized to the glucocorticoid cortisol by the microsomal CYP11B2. CYP21 can also metabolize progesterone to 11-deoxycorticosterone, which CYP11B2 converts to the glucocorticoid corticosterone, although this pathway is secondary in humans (although the principal pathway in rodents). (C) In the zona reticularis, CYP17 hydroxylates pregnenolone to 17-hydroxypregnenolone, and then DHEA. DHEA is the major steroid product of the reticularis, with sulfated DHEA (DHEA-S), androstenedione, and testosterone serving as only minor steroidogenic products.

Figure 3

Phylogenetic analysis of vertebrate mitochondrial CYP evolution. Homologs of the seven human mitochondrial CYPs (CYP11A1, CYP11B2, CYP11B3, CYP24A1, CYP27A1, CYP27B1, and CYP27C1) were identified using the nucleotide BLAST tool (41). Organisms chosen for search included the fully sequenced chicken (Gallus gallus), zebrafish (Danio rerio), and lancelet (Branchiostoma floridae), as they represented divergent vertebrate sequences (chicken vs. human vs. zebrafish) that could be compared with a non-vertebrate chordate (lancelet). The genomes of the Western clawed frog (Xenopus tropicalis) and anole lizard (Anolis carolinesis) were also searched, but sequences from these organisms were not included in the analysis as surprisingly no homologs of CYP11B1 or CYP11B2 were found (data not shown). Nucleotide sequences were aligned using MUSCLE (42), and phylogenetic trees constructed using PHyML (43).

Figure 4

Mitochondrial cytochrome P450 electron transport chain. The membrane-bound flavin-containing ferredoxin reductase (FDXR) accepts two electrons from NADPH, yielding NADP⁺. These electrons are passed to the iron–sulfur cluster of ferredoxin (FDX), which donates the electrons to the heme prosthetic group of the mitochondrial cytochrome P450 (CYP), which uses protons and molecular oxygen to hydroxylate its target substrate (R–H) yielding the final hydroxylated product (R–OH) and water.

Figure 5

Mitochondrial cholesterol transport and metabolism machinery. The mitochondrial cholesterol import and metabolism machinery are shown, demarcated by red (transduceosome) and blue (metabolon) dashed lines, respectively. The transduceosome contains cytoplasmic (StAR, ACBD3, DBI, and PRKARI; colored blue), OMM (VDAC and TSPO; colored green), and IMM (ATAD3A; colored red) components which assemble in response to hormonal stimulation and transduce the resultant cAMP signal to the mitochondria for cholesterol import. It is important to note, however, that molecular details of cholesterol import are still lacking. Once cholesterol is imported into the mitochondria, the IMM metabolon (CYP11A1, FDX, and FDXR; colored red) metabolizes cholesterol to pregnenolone, the precursor to all other steroids, including adrenal glucocorticoids and mineralocorticoids.