University of Texas Southwestern Medical Center Dallas, Texas 75235
Address all correspondence and requests for reprints to: Dr. William Kovacs, Division of Endocrinology, University of Texas, Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas 75235.
ACTH, the principal hormonal regulator of adrenal steroidogenesis, exerts its effects on the cells of the adrenal cortex by its interaction with a specific receptor resident in the cell membrane. This receptor is a member of a family of G protein-coupled receptors that have a common structure of seven membrane-spanning domains and are distinguished by their specificity for different ligands derived from the proopiomelanocortin precursor molecule (1). The ACTH receptor, designated melanocortin receptor 2 (MC2R), is encoded by a gene on the short arm of chromosome 18 (2). The gene does not contain introns within the coding sequence, but the transcriptional start site is in a small exon separated from the coding exon by approximately 18 kb of intervening sequence (3). The upstream promoter region contains response elements for steroidogenic factor 1 as well as several putative cAMP response elements (4, 5). In transient transfection experiments, the promoter has been demonstrated to be responsive to cAMP. Expression of the gene is increased by ACTH itself and the apparent half-life of the MC2R mRNA is increased by the action of the hormone (6, 7).
In this issue of the JCEM, Slawik et al. (8) report the identification of a relatively common polymorphism in the promoter of the MC2R gene. The polymorphism was first noted in a human genomic DNA clone from a genetic repository in Germany. This clone was found to have a cytosine instead of a thymine at a position 2 bp upstream of the transcriptional start site of the gene. The polymorphism does not involve any of the known or putative binding sites for regulatory transcription factors. When the authors surveyed a cohort of 1266 men from southern Germany, they found that almost 20% of the men had at least one copy of the variant MC2R gene.
This is the point at which molecular genetics and clinical physiology (or pathophysiology) frequently fail to meet. In this instance, however, a careful analysis of the phenotypic correlates of the genetic polymorphism has provided us with one of the first insights into a genetic basis for variation in hormonal responsiveness among apparently normal individuals.
The consequences of mutations in the coding region of the MC2R gene have been explored in some detail in subjects with a disorder called familial glucocorticoid deficiency (FGD) (9). This rare syndrome was identified over 45 yr ago as a form of hereditary Addisons disease (10). Individuals with the disorder exhibit signs and symptoms of primary adrenal insufficiency but without evidence of mineralocorticoid lack. Basal plasma cortisol levels are usually low or even undetectable and plasma ACTH levels are markedly elevated. Responses to exogenous ACTH are deficient. The first mutation in the MC2R gene was identified in a person with FGD shortly after the cloning of the receptor, and over 15 different mutations have been described to date; all are in the coding region of the gene and therefore all predict altered structure of the MC2R protein (9). Functional defects in some of these mutations have been characterized using transient transfection systems. However, not all kindreds with FGD have demonstrable defects in the coding region of MC2R, and postreceptor abnormalities, alterations in other regions of the MC2R gene (including the promoter), and alterations of other genes affecting ACTH signaling have all been postulated but none have been observed. Genetic variation in the region of the MC2R gene has been found; six single nucleotide polymorphisms are listed in the National Center for Biotechnology Informations single nucleotide polymorphism database, but the C/T polymorphism at position 2 has not been previously noted.
Because the sequence polymorphism noted in the MC2R gene in this German population was in the very proximal promoter region, and because it was known that the spontaneous mutations in MC2R dramatically affect adrenal responsiveness to ACTH, the authors postulated that the single-base change in the MC2R promoter might influence the level of expression of the variant allele and might consequently impact on either basal or stimulated adrenal steroid production. In transient transfection experiments in vitro, the variant promoter activity appeared lower than that of the "normal" promoter. Most remarkably, on testing in vivo, men with the variant MC2R gene exhibited diminished responsiveness to infused ACTH and were found to have higher ACTH/cortisol ratios in response to injection of CRH. Basal levels of plasma ACTH, cortisol, and dehydroepiandrosterone sulfate as well as urinary free cortisol and cortisone were indistinguishable among the various genotypes.
This demonstrated correlation between genotype and phenotype is an impressive achievement, especially because we can postulate some mechanisms to account for it; the authors favor a formulation in which diminished activity of the CCC promoter results in diminished expression of MC2R in adrenal cells, with consequent diminished responsiveness to ACTH. Nevertheless, it remains trite but true that this new work raises more questions than it answers. Is there a clinical significance to the findings? Are the CCC homozygotes or the CTC/CCC heterozygotes more susceptible to stresses than the CTC/CTC individuals? Or is more exuberant adrenal responsiveness unnecessary or even maladaptive under some circumstances? Do the demonstrated differences in adrenal responsiveness occur only at levels of hypothalamic-pituitary-adrenal (HPA) activity during which the decrement is physiologically insignificant?
Like most interesting experimental observations, these data immediately suggest some topics for further exploration. How do the individuals homozygous for the CCC MC2R gene respond to insulin-induced hypoglycemic challenge to the HPA axis? How do they respond to standard psychological stress tests? Do they have any degree of adrenal hyperplasia observable on imaging studies? Most importantly, is the CCC polymorphism associated with altered responsiveness to the stress of catastrophic illness, trauma, or major surgery, and are discernible differences in clinical outcomes observable in individuals with one or two copies of the variant promoter sequence?
The clinical relevance of such findings will likely be a complex problem to dissect, involving multiple genetic and physiological influences. From the strictly endocrinological standpoint, one could imagine that genetic differences at all levels of the HPA axis, including pituitary responses to CRH, maximal steroidogenic enzymatic activity of the adrenal, and metabolic clearance (or regeneration!) of cortisol by peripheral enzymatic systems, as well as ACTH responsiveness of the adrenal might all contribute to the overall response of the system to a major physiological stress.
The kind of phenotype/genotype correlation reported in this paper is really only a beginning, but as studies like those of Slawik et al. (8) reveal details of normal variation in the production, action, and metabolism of hormones, we can envision progress toward a sort of endocrine pharmaco-genomics, in which our understanding of the bases for individual differences in hormone responsiveness allows for therapeutic interventions more precisely tailored for individual patients and for their specific clinical situations.
Footnotes
Abbreviations: FGD, Familial glucocorticoid deficiency; HPA, hypothalamic-pituitary-adrenal; MC2R, melanocortin receptor 2.
Received May 13, 2004.
Accepted May 20, 2004.
References
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