Inactivating mutations of the pro-opiomelanocortin (POMC) gene in both mice and humans leads to hyperphagia and obesity. To further examine the mechanisms whereby POMC-deficiency leads to disordered energy homeostasis, we have generated mice lacking all POMC-derived peptides. Consistent with a previously reported model, Pomc ؊/؊ mice were obese and hyperphagic. They also showed reduced resting oxygen consumption associated with lowered serum levels of thyroxine. Hypothalami from Pomc ؊/؊ mice showed markedly increased expression of melanin-concentrating hormone mRNA in the lateral hypothalamus, but expression of neuropeptide Y mRNA in the arcuate nucleus was not altered. Provision of a 45% fat diet increased energy intake and body weight in both Pomc ؊/؊ and Pomc ؉/؊ mice. The effects of leptin on food intake and body weight were blunted in obese Pomc ؊/؊ mice whereas nonobese Pomc ؊/؊ mice were sensitive to leptin. Surprisingly, we found that Pomc ؊/؊ mice maintained their acute anorectic response to peptide-YY3-36 (PYY3-36). However, 7 days of PYY3-36 administration had no effect on cumulative food intake or body weight in wild-type or Pomc ؊/؊ mice. Thus, POMC peptides seem to be necessary for the normal response of energy balance to high-fat feeding, but not for the acute anorectic effect of PYY3-36 or full effects of leptin on feeding. The finding that the loss of only one copy of the Pomc gene is sufficient to render mice susceptible to the effects of high fat feeding emphasizes the potential importance of this locus as a site for gene-environment interactions predisposing to obesity. H ypothalamic neurons expressing pro-opiomelanocortin (POMC) are involved in the maintenance of energy homeostasis through the integration of a number of peripheral and central signals related to energy status (1-3). POMC is highly expressed in neuronal cell bodies of the arcuate nucleus, with POMC-expressing neurons innervating other hypothalamic regions known to regulate feeding behavior, including the paraventricular nucleus, lateral hypothalamus, and dorsomedial hypothalamic nucleus (DMH) (4). Pharmacological and genetic studies have revealed that POMCderived melanocortin peptides or synthetic agonists suppress feeding through activation of the melanocortin 4-receptor (MC4R) whereas the endogenous antagonist agouti-related protein (AgRP) or synthetic antagonists stimulate food intake (5).Approximately 40% of POMC-expressing neurons colocalize with the long isoform of the leptin receptor (6). Leptin activates POMC neurons directly and indirectly, through hyperpolarization of neuropeptide Y (NPY)͞AgRP neurons, thereby reducing their tonic inhibitory ␥-aminobutyric acid (GABA) input to POMC neurons (1). Recent data suggest that the gut peptide PYY 3-36 (peptide-YY 3-36 ) reduces food intake, at least in part, through direct hyperpolarization of NPY͞AgRP neurons and subsequent disinhibition of POMC neurons (3).Given the critical role that hypothalamic POMC plays in the regulation of energy balance, it is not surprising that m...

Proopiomelanocortin (POMC) can be processed to ACTH and melanocortin peptides. However, processing is incomplete in some tissues, leading to POMC precursor release from cells. This study examined POMC processing in human skin and the effect of POMC on the melanocortin-1 receptor (MC-1R) and melanocyte regulation. POMC was secreted by both human epidermal keratinocytes (from 5 healthy donors) and matched epidermal melanocytes in culture. Much lower levels of α-MSH were secreted and only by the keratinocytes. Neither cell type released ACTH. Cell extracts contained significantly more ACTH than POMC, and α-MSH was detected only in keratinocytes. Nevertheless, the POMC processing components, prohormone convertases 1, 2 and regulatory protein 7B2, were detected in melanocytes and keratinocytes. In contrast, hair follicle melanocytes secreted both POMC and α-MSH, and this was enhanced in response to corticotrophin-releasing hormone (CRH) acting primarily through the CRH receptor 1. In cells stably transfected with the MC-1R, POMC stimulated cAMP, albeit with a lower potency than ACTH, α-MSH, and β-MSH. POMC also increased melanogenesis and dendricity in human pigment cells. This release of POMC from skin cells and its functional activity at the MC-1R highlight the importance of POMC processing as a key regulatory event in the skin.-Rousseau, K., Kauser, S., Pritchard, L. E., Warhurst, A., Oliver, R. L., Slominski, A., Wei, E. T., Thody, A. J., Tobin, D. J., White, A. Proopiomelanocortin (POMC), the ACTH/melanocortin precursor, is secreted by human epidermal keratinocytes and melanocytes and stimulates melanogenesis.

The coding sequence for human serum transferrin was assembled from restriction fragments derived from a full-length cDNA clone isolated from a human liver cDNA library. The assembled clone was inserted into the expression vector pNUT and stably transfected into transformed baby hamster kidney (BHK) cells, leading to secretion of up to 125 mg/L recombinant protein into the tissue culture medium. As judged by mobility on NaDodSO4-PAGE, immunoreactivity, spectral properties (indicative of correct folding and iron binding), and the ability to bind to receptors on a human cell line, initial studies showed that the recombinant transferrin, is identical to three commercial human serum transferrin samples. Electrospray mass spectrometry (ESMS), anion-exchange chromatography, and urea gel analysis showed that the recombinant protein has an extremely complex carbohydrate pattern with 16 separate masses ranging from 78,833 to 80,802 daltons. Mutation of the two asparagine carbohydrate linkage sites to aspartic acid residues led to the expression and secretion of up to 25 mg/L nonglycosylated transferrin. ESMS, anion-exchange chromatography, and urea gel analysis showed a single molecular species that was consistent with the expected theoretical mass of 75,143 daltons. In equilibrium binding experiments, the nonglycosylated mutant bound to HeLa S3 cells with the same avidity and to the same extent as the glycosylated protein and the three commercial samples. These studies demonstrate conclusively that carbohydrate has no role in this function.