Elsevier

Archives of Medical Research

Volume 32, Issue 6, November–December 2001, Pages 576-586
Archives of Medical Research

Review article
Steroidogenic Acute Regulatory Protein: An Update on Its Regulation and Mechanism of Action

https://doi.org/10.1016/S0188-4409(01)00338-1Get rights and content

Abstract

Steroidogenic acute regulatory (StAR) protein controls the rate-limiting step in steroidogenesis: the transport of cholesterol from the outer to the inner mitochondrial membrane. Early studies indicated that rate of transcription of the StAR gene is a primary determinant of steroidogenesis. The transcription factors that govern basal and cAMP-dependent StAR expression are reviewed, as are new findings regarding chromatin modifications associated with activation of the StAR promoter. Molecular genetic studies of congenital lipoid adrenal hyperplasia, a rare disease caused by mutations in the StAR gene, and structure-function studies defined two major domains within the StAR protein, the N-terminal mitochondrial targeting sequence and the C-terminal StAR-related lipid transfer (START) domain, which promotes the translocation of cholesterol between the two mitochondrial membranes. Several models of StAR's mechanism of action have been proposed based on a combination of structure/function studies or on the crystal structure of a related START domain. The models—intermembrane shuttle hypothesis, and cholesterol desorption hypothesis—are discussed with respect to the known biochemical and biophysical events associated with steroidogenesis and the structure of StAR.

Introduction

Movement of cholesterol from the outer to the inner mitochondrial membrane is the rate-limiting step in steroid hormone synthesis. Steroidogenic acute regulatory (StAR) protein promotes the transfer of cholesterol across the aqueous space separating the cholesterol-rich outer mitochondrial membrane and cholesterol-poor inner mitochondrial membrane. In turn, cytochrome P450scc and its associated electron transport proteins, which reside on the matrix side of the inner mitochondrial membrane, convert cholesterol into pregnenolone. Evidence for a labile protein involved in steroidogenesis was obtained nearly 40 years ago when it was discovered that inhibitors of protein synthesis block hormonal induction of steroidogenesis 1, 2. Subsequent studies demonstrated that this block in steroidogenesis occurs prior to the formation of pregnenolone; more specifically, these inhibitors prevented the movement of cholesterol from the outer to inner mitochondrial membrane 1, 3. Moreover, disruption of the mitochondrial membranes increased steroidogenesis and addition of protein synthesis inhibitors failed to prevent this increase, suggesting that cholesterol access to side chain cleavage system was restricted in intact mitochondria (4). In contrast to cholesterol, soluble side-chain hydroxylated sterols stimulated high levels of steroid synthesis even in the presence of protein synthesis inhibitors and absence of tropic hormone stimulation 1, 2. Collectively, these observations suggested the existence of a tropic hormone-regulated protein that augments the movement of cholesterol from the outer to the inner mitochondrial membrane.

More than 20 years passed from the recognition that a protein was involved in hormonal-induced steroidogenesis before Orme-Johnston and colleagues (5) identified a group of mitochondrial 30 kDa phosphoproteins that appeared in adrenal cells stimulated with ACTH and gonadal cells stimulated with LH; their appearance could be blocked by protein synthesis inhibitors. Identical proteins were found in dibutyryl-cAMP-stimulated MA-10 mouse Leydig cells by Stocco and colleagues [see review (3)]. The 30 kDa proteins were shown to be derived from a 37 kDa precursor synthesized in the cytoplasm and then imported into mitochondria and processed into the 30 kDa forms. Purification of the 30 kDa protein from MA-10 cells and amino acid sequence analysis allowed the cloning of its cognate cDNA and the identification of a novel protein in the mouse (6) and human (7) named the steroidogenic acute regulatory protein (StAR).

StAR's role in hormonal-regulated steroid biosynthesis has since been confirmed by many investigators. Indeed, StAR's role in cholesterol movement has been the subject of several reviews, which can be examined for a more comprehensive summary of the studies performed to date 1, 2, 8, 9, 10, 11, 12, 13. In this article, we highlight the role molecular genetics has played in understanding the function of the StAR protein and recent observations regarding both transcriptional regulation of the StAR gene and posttranslational modification of the StAR protein. We also critically analyze the recently proposed models that explain how StAR acts.

Section snippets

StAR: Molecular Genetic Analysis of Congenital Lipoid Adrenal Hyperplasia

Molecular genetic analysis established the role of the StAR protein in intracellular cholesterol trafficking when it was discovered that congenital lipoid adrenal hyperplasia results from mutations in the StAR gene (14). Congenital lipoid adrenal hyperplasia is the most severe form of congenital adrenal hyperplasia characterized by a marked impairment of the biosynthesis of all adrenal and gonadal steroid hormones. The clinical phenotype of this disease includes the onset of profound

Regulation of StAR Expression and Activity: Transcription and Posttranslational Modifications

StAR was originally identified as a phosphoprotein and comparison of StAR protein sequences from multiple species indicated the presence of two consensus protein kinase A (PKA) motifs located at serine 57 and serine 195, both of which can be phosphorylated under in vitro and in vivo protocols (25). Sequence analysis also indicated that the protein contains putative phosphorylation sites for protein kinase C (PKC), CAM II kinase, creatine kinase, and P34CDC2 kinase. Recently, Calle et al. (26)

Transcriptional Regulation of StAR Expression

The study of the transcriptional regulation of StAR gene expression has garnered a significant amount of interest with the discovery that the presence of StAR transcripts in adrenal and gonadal cells is directly correlated with steroidogenesis 9, 28, 29. In the gonads and adrenal, tropic hormones stimulate steroid synthesis predominately through the cAMP-dependent pathway. Increases in intracellular cAMP levels are accompanied by rapid increases in StAR gene transcription and subsequently mRNA

Identification of a StAR-Related Lipid-Transfer (START) Domain

Protein sequence analysis revealed the presence of StAR homologs. MLN64 was the first gene recognized that exhibits homology to the StAR protein (67). The portion of the MLN64 sequence exhibiting homology to StAR was the carboxy-terminus, the essential portion of StAR that was found to stimulate steroidogenesis when co-transfected with the cholesterol side-chain cleavage enzyme system into COS-1 cells (68). Western blot analysis revealed that MLN64 fragments representing domains of MLN64 most

Structural Characterization of StAR and MLN64

Miller and colleagues characterized the StAR protein physical characteristics under different conditions (pH, heat, etc). These authors propose that StAR undergoes conformational changes in response to a change in pH, ultimately resulting in a loss of tertiary structure with no loss of secondary structure, or the formation of a molten globule (71). Molten globules are dynamic protein structures lacking fixed interactions that constrain the folded polypeptide chain organization. Because of the

Models of StAR Function

The presence of the mitochondrial targeting motif and cleavage sites for mitochondrial proteases within the rodent StAR protein, as well as the localization of StAR protein in putative contact sites between the outer and inner mitochondrial membranes, led to an initial model that suggested that StAR formed contacts sites between the mitochondrial membranes through which cholesterol could flow down a chemical gradient 9, 78. Unexpectedly, StAR mutant proteins lacking the first 62 amino acids

Conclusions

Because of the inherent difficulties in studying the transport of a small hydrophobic molecule such as cholesterol, progress in the field of intracellular lipid trafficking has been slow, as evidenced by the nearly 30 years between the first evidence for StAR's involvement in cholesterol metabolism and its subsequent discovery. Conversely, insight into the structural characterization of StAR and its physiologic role in steroidogenesis has advanced quickly, to a great extent due to the discovery

Acknowledgements

This work was supported by National Institutes of Health grant HD06274 to JFS.

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