Oleylethanolamide (OEA) is a naturally occurring lipid that regulates satiety and body weight. Although structurally related to the endogenous cannabinoid anandamide, OEA does not bind to cannabinoid receptors and its molecular targets have not been defined. Here we show that OEA binds with high affinity to the peroxisome-proliferator-activated receptor-alpha (PPAR-alpha), a nuclear receptor that regulates several aspects of lipid metabolism. Administration of OEA produces satiety and reduces body weight gain in wild-type mice, but not in mice deficient in PPAR-alpha. Two distinct PPAR-alpha agonists have similar effects that are also contingent on PPAR-alpha expression, whereas potent and selective agonists for PPAR-gamma and PPAR-beta/delta are ineffective. In the small intestine of wild-type but not PPAR-alpha-null mice, OEA regulates the expression of several PPAR-alpha target genes: it initiates the transcription of proteins involved in lipid metabolism and represses inducible nitric oxide synthase, an enzyme that may contribute to feeding stimulation. Our results, which show that OEA induces satiety by activating PPAR-alpha, identify an unexpected role for this nuclear receptor in regulating behaviour, and raise possibilities for the treatment of eating disorders.

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Summary3′-Uridylylation of RNA is emerging as a phylogenetically widespread phenomenon involved in processing events as diverse as uridine insertion/deletion RNA editing in mitochondria of trypanosomes and small nuclear RNA maturation in humans. This reaction is catalyzed by terminal uridylyltransferases (TUTases), which are template-independent RNA nucleotidyltransferases that specifically recognize UTP and belong to a large enzyme superfamily typified by DNA polymerase β. Multiple TUTases, recently identified in trypanosomes, as well as a U6 snRNA-specific TUTase enzyme in humans, are highly divergent at the protein sequence level. However, they all possess conserved catalytic and UTP recognition domains, often accompanied by various auxiliary modules present at the termini or between conserved domains. Here we report identification, structural and biochemical analyses of a novel trypanosomal TUTase, TbTUT4, which represents a minimal catalytically active RNA uridylyltransferase. The TbTUT4 consists of only two domains that define the catalytic center at the bottom of the nucleoside triphosphate and RNA substrate binding cleft. The 2.0 Å crystal structure reveals two significantly different conformations of this TUTase: one molecule is in a relatively open apo conformation, whereas the other displays a more compact TUTase-UTP complex. A single nucleoside triphosphate is bound in the active site by a complex network of interactions between amino acid residues, a magnesium ion and highly ordered water molecules with the UTP's base, ribose and phosphate moieties. The structure-guided mutagenesis and cross-linking studies define the amino acids essential for catalysis, uracil base recognition, ribose binding and phosphate coordination by uridylyltransferases. In addition, the cluster of positively charged residues involved in RNA binding is identified. We also report a 2.4 Å crystal structure of TbTUT4 with the bound 2′ deoxyribonucleoside, which provides the structural basis of the enzyme's preference toward ribonucleotides.

A nnexins are structurally divided into a highly conserved core domain and a variable N-terminal domain. The core domain mediates the calcium-dependent phospholipid binding of annexins, whereas the N-terminal domain, which is unique in sequence and length for each member of this protein family, is responsible for the specificity among the different members. Annexin 1 has been shown to possess membrane aggregation and fusion activity in the presence of calcium in vitro. Due to the high sequence homology of the core domain among different annexins, the property of membrane aggregation has been attributed to the N-terminal domain. For instance, a chimera protein comprising the core of annexin 5, which by itself does not exhibit membrane aggregation properties, and the N-terminal domain of annexin 1 is able to induce membrane aggregation. Numerous three-dimensional structures of annexins have been solved using x-ray crystallography, however, none reveal the tertiary structure of an N-terminal domain or its interaction with the core domain. We have solved the x-ray structure of full-length annexin 1 with an N-terminal domain comprising 42 amino acids in the absence of calcium ions at 1.8 Å resolution. Residues 2-26 of the N-terminal domain exhibit a mainly α-helical conformation with a kink at residue 17. Helix NA (residues 2 to 16) inserts into repeat III of the core domain thereby replacing the old helix D. Helix D on the other hand unfolds into an extended loop that forms a flap over the top of helix NA of the N-terminal domain. As a result, the type II calcium-binding site located in core repeat III is destroyed because the "capping residue" for calcium ion coordination is not in the proper conformation/location any more. Also presented in this article is the structure of full-length annexin 1 in the presence of 1 mM CaCl 2 . The structure of full-length annexin 1 in the absence of calcium ions is thought to represent the "inactive" form of the protein. We provide a model for the annexin 1-induced membrane aggregation and discuss it in light of the literature published to date.