Identification of nesfatin-1 as a satiety molecule in the hypothalamus

Abstract

The brain hypothalamus contains certain secreted molecules that are important in regulating feeding behaviour. Here we show that nesfatin, corresponding to NEFA/nucleobindin2 (NUCB2), a secreted protein of unknown function, is expressed in the appetite-control hypothalamic nuclei in rats. Intracerebroventricular (i.c.v.) injection of NUCB2 reduces feeding. Rat cerebrospinal fluid contains nesfatin-1, an amino-terminal fragment derived from NUCB2, and its expression is decreased in the hypothalamic paraventricular nucleus under starved conditions. I.c.v. injection of nesfatin-1 decreases food intake in a dose-dependent manner, whereas injection of an antibody neutralizing nesfatin-1 stimulates appetite. In contrast, i.c.v. injection of other possible fragments processed from NUCB2 does not promote satiety, and conversion of NUCB2 to nesfatin-1 is necessary to induce feeding suppression. Chronic i.c.v. injection of nesfatin-1 reduces body weight, whereas rats gain body weight after chronic i.c.v. injection of antisense morpholino oligonucleotide against the gene encoding NUCB2. Nesfatin-1-induced anorexia occurs in Zucker rats with a leptin receptor mutation, and an anti-nesfatin-1 antibody does not block leptin-induced anorexia. In contrast, central injection of alpha-melanocyte-stimulating hormone elevates NUCB2 gene expression in the paraventricular nucleus, and satiety by nesfatin-1 is abolished by an antagonist of the melanocortin-3/4 receptor. We identify nesfatin-1 as a satiety molecule that is associated with melanocortin signalling in the hypothalamus.

Expanding roles of NUCB2/nesfatin-1 in neuroendocrine regulation

“…Indeed, by the generation of specific antibodies and implementation of immunohistochemical (IHC) analyses and immunoassays, nesfatin-1 was shown to be produced in the hypothalamus and present in the cerebrospinal fluid (index of this being a secreted neuropeptide). Co-localization of nesfatin-1 and PC3/1 and PC2 suggested that the latter are involved in the cleavage of nesfatin-1 from the NUCB2 precursor (Oh et al 2006). …”

“…Among the clones identified to be regulated by PPAR-g, one was highlighted as i) having a signal peptide (index of secreted proteins), ii) being produced by neuronal-(medulloblastoma HTB185) and adipocyte-(3T3-L1) derived cell lines, and iii) displaying profound stimulated expression following troglitazone (Oh et al 2006). This target was shown to correspond to the gene encoding nucleobindin 2 (NUCB2), also termed NEFA (for DNA binding/EF-hand/acidic protein).…”

“…In the search for new appetite-controlling signals, Oh et al (2006) reported a series of analyses based on a subtraction cloning strategy in SQ-5 cells (derived from lung carcinoma, which expresses leptin and leptin receptors (Tsuchiya et al 1999)), following stimulation with the PPAR-g ligand, troglitazone. Among the clones identified to be regulated by PPAR-g, one was highlighted as i) having a signal peptide (index of secreted proteins), ii) being produced by neuronal-(medulloblastoma HTB185) and adipocyte-(3T3-L1) derived cell lines, and iii) displaying profound stimulated expression following troglitazone (Oh et al 2006).…”

“…NUCB2 and its closely related NUCB1 are known to be secreted proteins (Miura et al 1992), but their functions remained largely unknown at that time. Based on the additional observations that NUCB2 is indeed expressed in key hypothalamic nuclei involved in body weight control (see below) and that, when injected into the brain, recombinant NUCB2 protein reduces food intake, this factor was termed nesfatin (for NEFA/NUCB2-encoded satiety-and fat-influencing proteins) (Oh et al 2006). NUCB2/nesfatin is composed of 396 amino acids, preceded by a 24-amino acid signal peptide ( Fig.…”

“…Structural analyses revealed the presence of several conserved cleavage recognition sites for prohormone convertases (PC) within rat NUCB2/nesfatin sequence, thus suggesting this to be a precursor that gives rise, by differential proteolytic processing, to several active peptides. The predicted (major) fragments of such processing were termed nesfatin-1 (spanning residues 1-82), nesfatin-2 (residues 85-163), and nesfatin-3 (residues 166-396) (Oh et al 2006). Indeed, by the generation of specific antibodies and implementation of immunohistochemical (IHC) analyses and immunoassays, nesfatin-1 was shown to be produced in the hypothalamus and present in the cerebrospinal fluid (index of this being a secreted neuropeptide).…”