Peroxisome Proliferator-activated Receptor Coactivator-1 (PGC-1 ) Coactivates the Cardiac-enriched Nuclear Receptors Estrogen-related Receptor- and - IDENTIFICATION OF NOVEL LEUCINE-RICH INTERACTION MOTIF

The transcriptional coactivator PPAR coactivator-1 (PGC-1 ) has been characterized as a broad regulator of cellular energy metabolism. Although PGC-1 functions through many transcription factors, the PGC-1 partners identified to date are unlikely to account for all of its biologic actions. The orphan nuclear receptor estrogen-related receptor (ERR ) was identified in a yeast two-hybrid screen of a cardiac cDNA library as a novel PGC-1 -binding protein. ERR was implicated previously in regulating the gene encoding medium-chain acyl-CoA dehydrogenase (MCAD), which catalyzes the initial step in mitochondrial fatty acid oxidation. The cardiac perinatal expression pattern of ERR paralleled that of PGC-1 and MCAD. Adenoviral-mediated ERR overexpression in primary neonatal cardiac mycoytes induced endogenous MCAD expression. Furthermore, PGC-1 enhanced the transactivation of reporter plasmids containing an estrogen response element or the MCAD gene promoter by ERR and the related isoform ERR . In vitro binding experiments demonstrated that ERR interacts with PGC-1 via its activation function-2 homology region. Mutagenesis studies revealed that the LXXLL motif at amino acid position 142–146 of PGC-1 (L2), necessary for PGC-1 interactions with other nuclear receptors, is not required for the PGC1 ERR interaction. Rather, ERR binds PGC-1 primarily through a Leu-rich motif at amino acids 209–213 (Leu-3) and utilizes additional LXXLL-containing domains as accessory binding sites. Thus, the PGC1 ERR interaction is distinct from that of other nuclear receptor PGC-1 partners, including PPAR , hepatocyte nuclear factor-4 , and estrogen receptor . These results identify ERR and ERR as novel PGC-1 interacting proteins, implicate ERR isoforms in the regulation of mitochondrial energy metabolism, and suggest a potential mechanism whereby PGC-1 selectively binds transcription factor partners. Cellular energy production is tightly linked to metabolic demand, which is, in turn, dictated by diverse developmental, physiologic, and environmental conditions. The capacity for cellular ATP production is controlled, in part, by the expression levels of nuclear genes involved in mitochondrial oxidative metabolism. Thus, tight regulation of cellular energy metabolism necessitates transduction of diverse signals related to cellular energy demands to the nucleus. Although numerous factors involved in the transcriptional regulation of metabolic gene expression have been identified, the precise pathways involved in the physiologic control of cellular energy metabolism have not been delineated. The recent discovery of PPAR coactivator-1 (PGC-1 ), PGC-1 , and the PGC-1-related protein, a family of inducible transcriptional coactivators responsive to selective physiological stimuli, have provided new insights into the link between extracellular events and the regulation of genes involved in energy metabolism. PGC-1 , the first member of this novel coactivator family to be identified, was initially characterized as a key regulator of thermogenesis in brown adipose tissue (BAT) and skeletal muscle via its coactivation of the adipose-enriched nuclear receptor, PPAR (1, 2). Subsequent studies have revealed a broader role for PGC-1 in a variety of cellular energy metabolic processes including mitochondrial biogenesis, mitochondrial fatty acid oxidation (FAO), and gluconeogenesis (2–6). The function of PGC-1 and PGC-1-related protein remain to be defined. PGC-1 is unique from the p160 and p300/cAMP response element-binding protein-binding protein classes of transcriptional coactivators in its tissue-restricted expression pattern, its developmental regulation, and its inducibility by specific physiological stimuli. PGC-1 is enriched in tissues reliant on oxidative metabolism for ATP generation (heart, skeletal muscle) or heat (BAT) but is also expressed in liver, brain, and kidney (1). Immediately after birth, PGC-1 expression increases in heart coincident with a shift from reliance on glycolysis to mitochondrial FAO as the chief energy source in the adult myocardium (4). PGC-1 expression is induced in adult skeletal muscle, BAT, and heart in response to stimuli that increase energy demands. For example, cold exposure leads to * This work was supported by National Institutes of Health Grants RO1 DK45416, RO1 HL58493, P30 DK56341, and P30 DK52574. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. ‡ Supported by National Institutes of Health National Research Service Award F32 HL10410. § To whom correspondence should be addressed: Center for Cardiovascular Research, Washington University School of Medicine, 660 S. Euclid Ave., Campus Box 8086, St. Louis, MO 63110. Tel.: 314-3628908; Fax: 314-362-0186; E-mail: [email protected]. 1 The abbreviations used are: PGC-1 , PPAR coactivator-1 ; BAT, brown adipose tissue; PPAR, peroxisome proliferator-activated receptor; FAO, fatty acid oxidation; MCAD, medium-chain acyl-CoA dehydrogenase; ERR, estrogen-related receptor; GFP, green fluorescent protein; COUP-TF, chicken ovalbumin upstream promoter transcription factor; GST, glutathione S-transferase; aa, amino acid; LBD, ligand binding domain; DBD, DNA binding domain; NRRE, nuclear receptor response element; MAPK, mitogen-activated protein kinase; NR, nuclear receptor; HNF-4 , hepatocyte nuclear factor-4 ; ER , estrogen receptor . THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol. 277, No. 43, Issue of October 25, pp. 40265–40274, 2002 © 2002 by The American Society for Biochemistry and Molecular Biology, Inc. Printed in U.S.A.