Growth/differentiation factor 15 (GDF15), also known as MIC-1, is a distant member of the transforming growth factor-β (TGF-β) superfamily and has been implicated in various biological functions, including cancer cachexia, renal and heart failure, atherosclerosis and metabolism. A connection between GDF15 and body-weight regulation was initially suggested on the basis of an observation that increasing GDF15 levels in serum correlated with weight loss in individuals with advanced prostate cancer. In animal models, overexpression of GDF15 leads to a lean phenotype, hypophagia and other improvements in metabolic parameters, suggesting that recombinant GDF15 protein could potentially be used in the treatment of obesity and type 2 diabetes. However, the signaling and mechanism of action of GDF15 are poorly understood owing to the absence of a clearly identified cognate receptor. Here we report that GDNF-family receptor α-like (GFRAL), an orphan member of the GFR-α family, is a high-affinity receptor for GDF15. GFRAL binds to GDF15 in vitro and is required for the metabolic actions of GDF15 with respect to body weight and food intake in vivo in mice. Gfral mice were refractory to the effects of recombinant human GDF15 on body-weight, food-intake and glucose parameters. Blocking the interaction between GDF15 and GFRAL with a monoclonal antibody prevented the metabolic effects of GDF15 in rats. Gfral mRNA is highly expressed in the area postrema of mouse, rat and monkey, in accordance with previous reports implicating this region of the brain in the metabolic actions of GDF15 (refs. 4,5,6). Together, our data demonstrate that GFRAL is a receptor for GDF15 that mediates the metabolic effects of GDF15.

Two new cloned human cDNAs encode paralogs of the 85-kDa cytosolic phospholipase A 2 (cPLA 2 ). We propose to call these cPLA 2 ␤ (114 kDa) and cPLA 2 ␥ (61 kDa), giving the name cPLA 2 ␣ to the well known 85-kDa enzyme. cPLA 2 ␤ mRNA is expressed more highly in cerebellum and pancreas and cPLA 2 ␥ more highly in cardiac and skeletal muscle. Sequence-tagged site mapping places cPLA 2 ␤ on chromosome 15 in a region near a phosphoinositol bisphosphate phosphatase. The mRNA for cPLA 2 ␤ is spliced only at a very low level, and Northern blots in 24 tissues show exclusively the unspliced form. cPLA 2 ␤ has much lower activity on 2-arachidonoyl-phosphatidylcholine liposomes than either of the other two enzymes. Its sequence contains a histidine motif characteristic of the catalytic center of caspase proteases of the apoptotic cascade but no region characteristic of the catalytic cysteine. Sequence-tagged site mapping places cPLA 2 ␥ on chromosome 19 near calmodulin. cPLA 2 ␥ lacks the C2 domain, which gives cPLA 2 ␣ its Ca 2؉ sensitivity, and accordingly cPLA 2 ␥ has no dependence upon calcium, although cPLA 2 ␤ does. cPLA 2 ␥ contains a prenyl group-binding site motif and appears to be largely membrane-bound. cPLA 2 ␣ residues activated by phosphorylation do not appear to be well conserved in either new enzyme. In contrast, all three previously known catalytic residues, as well as one additional essential arginine, Arg-566 in cPLA 2 ␣, are conserved in both new enzyme sequences. Mutagenesis shows strong dependence on these residues for catalytic activity of all three enzymes.Enzymatic breakdown of glycerophospholipids is carried out by numerous pathways, with the production of various bioactive lipids and fatty acids (1-8). Diverse phospholipase A 2 (PLA 2 ) 1 enzymes hydrolyze the sn-2 bond of phospholipids, releasing lysophospholipids and fatty acids (9 -11). Often the released fatty acid is arachidonic acid, whose further metabolism gives rise to several types of bioactive lipids known as eicosanoids, many of which mediate inflammation. Thus PLA 2 enzymes initiate the production of inflammatory mediators (12, 13), and for that reason they have become targets for the development of anti-inflammatory therapies.The 85-kDa cytosolic phospholipase A 2 (reviewed in Refs. 12-15) has attracted special interest because it is the only one of numerous PLA 2 s that selectively releases arachidonic acid over other fatty acids (16,17). Recent results with transgenic mice ablated for this enzyme (18) have demonstrated its role in allergy and parturition. Several functional regions have been identified within its amino acid sequence, including the C2 or calcium and lipid binding region similar to the C2 regions of other proteins (19) such as the calcium-dependent protein kinase C enzymes; several serine residues capable of activation through phosphorylation (20 -22); a hydrophilic region that may have a structural role (13); and several residues essential for catalysis, identified by mutagenesis (23-25).The cDNA for the 85-kDa cP...

The imidazoline-1 receptor (IR1) is considered a novel target for drug discovery. Toward cloning an IR1, a truncated cDNA clone was isolated from a human hippocampal lambda gt11 cDNA expression library by relying on the selectivity of two antisera directed against candidate IR proteins. Amplification reactions were performed to extend the 5' and 3' ends of this cDNA, followed by end-to-end PCR and conventional cloning. The resultant 5131-basepair molecule, designated imidazoline receptor-antisera-selected (IRAS) cDNA, was shown to encode a 1504-amino acid protein (IRAS-1). No relation exists between the amino acid sequence of IRAS-1 and proteins known to bind imidazolines (e.g., it is not an alpha2-adrenoceptor or monoamine oxidase subtype). However, certain sequences within IRAS-1 are consistent with signaling motifs found in cytokine receptors, as previously suggested for an IR1. An acidic region in IRAS-1 having an amino acid sequence nearly identical to that of ryanodine receptors led to the demonstration that ruthenium red, a dye that binds the acidic region in ryanodine receptors, also stained IRAS-1 as a 167-kD band on SDS gels and inhibited radioligand binding of native I1 sites in untransfected PC-12 cells (a source of authentic I1 binding sites). Two epitope-selective antisera were also generated against IRAS-1, and both reacted with the same 167-kD band on Western blots. In a host-cell-specific manner, transfection of IRAS cDNA into Chinese hamster ovary cells led to high-affinity I1 binding sites by criteria of nanomolar affinity for moxonidine and rilmenidine. Thus, IRAS-1 is the first protein discovered with characteristics of an IR1.

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Cytosolic phospholipase A 2 (cPLA 2 ) hydrolyzes the sn-2-acyl ester bond of phospholipids and shows a preference for arachidonic acid-containing substrates. We found previously that Ser-228 is essential for enzyme activity and is likely to function as a nucleophile in the catalytic center of the enzyme (