Conserved amino acids in the ligand-binding and tau(i) domains of the peroxisome proliferator-activated receptor alpha are necessary for heterodimerization with RXR.
The peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor superfamily. These ligand-activated transcription factors are implicated in the regulation of lipid metabolism and adipocyte differentiation and in the regulation of anti-inflammatory processes. In order to bind to DNA and activate transcription PPAR requires the formation of heterodimers with the retinoid X receptor (RXR). We have previously reported that replacement of a single leucine by an arginine at position 433 of hPPAR alpha (L433R), located in a highly conserved region of the ninth heptad repeat of a leucine-zipper-like motif in the ligand binding domain, abolished heterodimerization of PPAR with RXR and hence its trans-activating capacity. The aim of our present work was to investigate if other conserved amino acids of the ligand binding domain are important for heterodimerization of PPAR with RXR. We found that conserved leucines, L370 and L391, in a leucine-zipper-like motif of hPPAR alpha, as well as a highly conserved aspartic acid (D304) in the tau(i) domain are necessary for heterodimerization with RXR. In contrast, mutations of non-conserved amino acids within the leucine-zipper-like motif do not affect PPAR:RXR heterodimerization. Surprisingly, we found that some mutants deficient in heterodimerization with RXR (hPPAR alpha-L370R and -L391R) were still functional on specific peroxisome proliferator-activator response elements (PPREs). Both mutants could trans-activate on a PPRE from the P450 cytochrome promoter CYP4A1, whereas only the hPPAR alpha-L391R mutant could trans-activate from the acyl-CoA oxidase PPRE (ACOA) and, when stimulated with the peroxisome proliferator Wy14643, also from the bifunctional enzyme PPRE. We therefore hypothesize either that: (i) these mutants might be able to heterodimerize with a protein other than RXR and the affinity for this novel partner may depend on the nature of the PPRE and to some degree on the choice of the activator, or alternatively; (ii) that additional nuclear proteins might compensate in vivo for the decreased binding of RXR to these mutant PPARs observed in vitro.
Gorla-Bajszczak
A
A
Division d'Endocrinologie et Diabétologie, Hôpital Cantonal Universitaire de Genève, Geneva, Switzerland.
Juge-Aubry
C
C
Pernin
A
A
Burger
A G
AG
Meier
C A
CA
D042962:Acyl-CoA Oxidase; D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000235:genetics / Q000378:metabolism; D017124:Conserved Sequence / Q000235:genetics / Q000502:physiology*; D042926:Cytochrome P-450 CYP4A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D007930:Leucine / Q000235:genetics / Q000378:metabolism; D016350:Leucine Zippers / Q000235:genetics; D008024:Ligands; D006899:Mixed Function Oxygenases / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011743:Pyrimidines / Q000494:pharmacology; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018168:Receptors, Retinoic Acid / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D047488:Retinoid X Receptors; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D008024:Ligands; D011743:Pyrimidines; D018160:Receptors, Cytoplasmic and Nuclear; D018168:Receptors, Retinoic Acid; D047488:Retinoid X Receptors; D015534:Trans-Activators; D014157:Transcription Factors; D001224:Aspartic Acid; C006253:pirinixic acid; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D010088:Oxidoreductases; D042926:Cytochrome P-450 CYP4A; D042962:Acyl-CoA Oxidase; D007930:Leucine