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Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine, Miami, Florida 33136
Address all correspondence and requests for reprints to: Kerry L. Burnstein, Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine, 1600 Northwest 10th Avenue, R-189, Miami, Florida 33136.
Analysis of individuals with hormone resistance has historically provided valuable clues to the physiologic actions of hormones as well as the cognate receptors. Hormone resistance or insensitivity disorders are characterized by limited or absent hormone responsiveness despite the presence of normal, or even elevated, levels of circulating hormone. In clinical cases of steroid hormone and nuclear receptor ligand resistance, the literature is replete with evidence that receptor mutations underlie the syndromes. Although the advent of molecular cloning has allowed extensive site-directed mutagenesis studies, examination of naturally occurring mutations continues to afford the opportunity for unanticipated and novel insights into ligand and receptor structure/function relationships. In this issue of Endocrinology, Agostini and Gurnell et al. (1) characterize two different, naturally occurring loss-of-function mutations in the peroxisome proliferator-activated receptor (PPAR) 
, a nuclear receptor. These PPAR mutant alleles were previously found to be associated with three human cases of PPAR insensitivity characterized by severe insulin resistance (2).
The nuclear receptor superfamily is a diverse group of ligand-activated transcription factors including steroid/retinoid/thyroid hormone receptors and other more recently identified members such as the PPARs 
, 
, and (reviewed in Refs. 3, 4, 5). A similar organization of functional domains required for DNA and ligand binding as well as for transcriptional activity typically characterize nuclear receptors. The highly conserved and centrally positioned DNA binding domains of nuclear receptors make contacts with specific DNA sequence elements that are associated with appro-priate target genes. The ligand binding domains, located in carboxyl-terminal regions, represents a major portion of these receptors and consist of 12 -helices. Helices 1–11 assemble in a three-layer structure that generates a large hydrophobic pocket for ligand occupancy. Helix 12 exhibits substantial mobility and usually adopts different orientations in the aporeceptor vs. ligand-bound receptor (reviewed in Ref. 6). In addition to ligand binding, this domain contains subregions for dimerization and interaction with cofactors/coregulators. Ligand binding induces a conformational change resulting in release of inhibitory coregulators termed corepressors and recruitment of coactivators (reviewed in Refs. 7 and 8). Coactivators and associated factors possess enzymatic activities including histone acetylases and thereby remodel local chromatin structure resulting in enhanced transcriptional activity. Corepressors recruit histone deactylases and factors with other enzymatic activities leading to chromatin compaction. Although steroid receptors bind to DNA as homodimers, a major and growing subgroup of nuclear receptors including thyroid hormone receptor, retinoid receptors, and PPARs bind DNA as heterodimeric complexes with a common partner, the retinoid X receptor.
Both PPAR mutants (P467L and V290M) characterized in Agostini and Gurnell et al. (1) localize to the ligand binding domain, are not activated by putative endogenous ligands (including a variety of unsaturated fatty acids, eicosanoids, and 15-deoxy
12,14-prostaglandin J2), and exhibit diminished responses to thiazolidinediones, a class of insulin-sensitizing drugs that serve as high affinity PPAR agonists. In addition, these receptors exert dominant negative repression of wild-type PPAR transcriptional activity (1, 2). Alterations that result in receptors with dominant negative activity are of particular interest as individuals heterozygous for the mutation may exhibit phenotypes approaching complete insensitivity. Indeed, subjects expressing these mutant PPAR alleles exhibit severe insulin resistance, type II diabetes mellitus, features of the human metabolic syndrome (dyslipidemia and hypertension), and partial lipodystrophy in agreement with known and emerging roles for PPAR in regulating insulin sensitivity, glucose metabolism, adipogenesis, and blood pressure (1, 2). Although PPAR mutations appear to be rare in severe insulin resistance (three mutations were found in 85 cases), the work by Agostini and Gurnell et al. (1) provides significant insight into PPAR function, as well as the mechanism of action of a new class of PPAR agonist.
Although the response to thiazolidinediones (e.g. rosiglitazone) was markedly impaired, a new class of high-affinity, tyrosine-based receptor agonists (exemplified by farglitazar) promoted wild-type levels of transcriptional activation by the PPAR (V290M and P467L) mutants (1). Farglitazar permitted release of the corepressor, silencing mediator of retinoic acid and thyroid hormone receptors (SMRT), and the subsequent binding of coactivators. Based on previous work showing that these mutations alter the position of helix 12 (9) and on the structure of a PPAR-SMRT peptide complex (10), the authors speculate that helix 12 of the mutant receptors is destabilized, favoring receptor interaction with SMRT and resulting in dominant negative activity. Consistent with this model, dominant negative activity of the P467L mutant is abolished by an additional mutation (L318A) that disrupts corepressor interaction. The structural basis for the response to farglitazar is proposed to be via improved ligand-mediated stabilization of helix 12, thereby favoring corepressor release, coactivator binding, and regulation of target gene expression (Fig. 1
).
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coregulator binding helices lead to the conclusion that corepressor dissociation is the limiting step in coactivator binding and the activation of certain nuclear receptors (11). Interestingly, corepressor release may not be as critical a step in the activation of wild-type PPAR. Helix 12 appears to be in an active conformation in the apo-receptor (12), and the receptor exhibits relatively high basal activity (1, 13), suggesting that coactivator may bind even in the absence of ligand. The PPAR mutations (V290M and P467L) appear to shift the receptor to a situation where, much like the case for many nuclear receptors, corepressor release is a critical prerequisite for activation. Deregulation of corepressor-nuclear receptor interactions underlies other human disorders. In the case of resistance to thyroid hormone, dominant negative thyroid hormone receptor ß mutants have been identified in receptor ligand binding domains. Notably, several of these mutants display defects in corepressor dissociation (for example see Refs. 14 and 15). Similarly, failure of corepressor release underlies certain leukemias (reviewed in Refs. 16 and 17). In acute promyelocytic leukemia, chromosomal translocations of RAR frequently result in fusions with the promyelocytic leukemia gene or with the promyelocytic leukemia zinc finger gene. Both SMRT and another corepressor, nuclear receptor corepressor, bind to the fusion proteins due to retention of the RAR-interacting regions. Both fusion proteins fail to release corepressors after addition of the RAR ligand retinoic acid; however, pharmacologic concentrations of retinoic acid can overcome the RAR/promyelocytic leukemia-corepressor interaction.
Aberrant corepressor interactions resulting from mutations, or fusion proteins generated by chromosomal translocations, may underlie the lack of or attenuated clinical response to nuclear receptor ligands. In the case of PPAR mutants, Agostini and Gurnell et al. (1) now show that new, higher-affinity agonists have the potential to subvert this proposed disease mechanism. This work also supports the continued value of using natural mutations to understand receptor structure, function, and pharmacology. Furthermore, under some circumstances the therapeutic targeting of corepressor release from nuclear receptors is a rational and exciting goal.
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Footnotes |
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Received January 4, 2004.
Accepted for publication January 6, 2004.
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References |
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. Endocrinology 145:1527–1538 associated with severe insulin resistance, diabetes mellitus and hypertension. Nature 402:880–883[Medline]
. Nature 415:813–817[Medline]
ligand-binding domain. J Biol Chem 278:7366–7373
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