Characterization of the Amino-terminal Activation Domain of Peroxisome Proliferator-activated Receptor α

Abstract

The transactivating function of the A/B region of mouse peroxisome proliferator-activated receptor alpha (PPARalpha; NR1C1) was characterized. The truncated version of PPARalpha lacking the A/B region had 60-70% lower transactivating function than full-length PPARalpha in both the presence and absence of the peroxisome proliferator ciprofibrate. When tethered to the yeast Gal4 DNA-binding domain, the A/B region exhibited the significant ligand-independent transactivating function, AF-1 activity. The first 44 amino acid residues were necessary for maximal transactivation, and the minimally essential region was further delimited to amino acids 15-44. This region is highly enriched with acidic residues, but mutational analyses showed that the protein structure, rather than the negative charge itself, was important for the AF-1 activity. An alpha-helical configuration was predicted for this region, and a CD spectrum analysis of the synthetic peptides showed that mutant sequences with higher AF-1 activity have higher helical contents and vice versa. The most active mutant, in which Met(31) was replaced with Leu, was approximately 5-fold more potent than the wild-type A/B region. These findings indicate that the AF-1 region of PPARalpha is an acidic activation domain and that the helix-forming property is implicated in the transactivating function.