RESUMEN
OBJECTIVE: Obesity is a major health problem associated with high morbidity and mortality. NSAID-activated gene (NAG-1) is a TGF-ß superfamily member reported to alter adipose tissue levels in mice. We investigated whether hNAG-1 acts as a regulator of adiposity and energy metabolism. DESIGN/SUBJECTS: hNAG-1 mice, ubiquitously expressing hNAG-1, were placed on a control or high-fat diet for 12 weeks. hNAG-1-expressing B16/F10 melanoma cells were used in a xenograft model to deliver hNAG-1 to obese C57BL/6 mice. RESULTS: As compared with wild-type littermates, transgenic hNAG-1 mice have less white fat and brown fat despite equivalent food intake, improved glucose tolerance, lower insulin levels and are resistant to dietary- and genetic-induced obesity. hNAG-1 mice are more metabolically active with higher energy expenditure. Obese C57BL/6 mice treated with hNAG-1-expressing xenografts show decreases in adipose tissue and serum insulin levels. hNAG-1 mice and obese mice treated with hNAG-1-expressing xenografts show increased thermogenic gene expression (UCP1, PGC1α, ECH1, Cox8b, Dio2, Cyc1, PGC1ß, PPARα, Elvol3) in brown adipose tissue (BAT) and increased expression of lipolytic genes (Adrb3, ATGL, HSL) in both white adipose tissue (WAT) and BAT, consistent with higher energy metabolism. CONCLUSION: hNAG-1 modulates metabolic activity by increasing the expression of key thermogenic and lipolytic genes in BAT and WAT. hNAG-1 appears to be a novel therapeutic target in preventing and treating obesity and insulin resistance.
Asunto(s)
Tejido Adiposo/metabolismo , Factor 15 de Diferenciación de Crecimiento/metabolismo , Lipólisis , Obesidad/prevención & control , Termogénesis , Tejido Adiposo/patología , Animales , Western Blotting , Dieta Alta en Grasa , Ingestión de Alimentos , Metabolismo Energético , Ensayo de Inmunoadsorción Enzimática , Humanos , Resistencia a la Insulina , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Estrés Oxidativo/efectos de los fármacos , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
NAG-1/GDF15 is a TGF- ß superfamily member with poorly characterized biological activity proposed to inhibit inflammatory cytokine production. Transgenic mice expressing human NAG-1/GDF15 (NAG-1 (Tg/Lox) ) are leaner with lower body weight and are resistant to chemically or genetically induced intestinal tumors. Because of the link between obesity, inflammation, and cancer, we examined whether these mice exhibit a reduced response to inflammatory stimuli. The NAG-1 (Tg/Lox) mice had a reduced inflammatory response to LPS based on the serum levels of cytokines KC, IL-6, MCP-1, and TNF α . In contrast to literature reports and our in vivo results, NAG-1 did not inhibit LPS-induced cytokine expression in vitro in RAW264.7 cells, mouse peritoneal macrophages, or mouse liver Kupffer cells, suggesting that NAG-1/GDF15 does not directly inhibit LPS-induced inflammatory cytokine production. However, NAG-1 (Tg/Lox) mice have less white adipose tissue, the major source of inflammatory adipokines including leptin. Basal and LPS-treated serum leptin and mRNA levels in the adipose tissue of NAG-1 (Tg/Lox) mice were lower than those in WT mice. We propose that the reduced white adipose tissue and reduced leptin expression may be responsible, in part, for the reduced inflammatory response to LPS and the decrease in intestinal tumors observed in NAG-1 (Tg/Lox) mice.
Asunto(s)
Tejido Adiposo Blanco/metabolismo , Factor 15 de Diferenciación de Crecimiento/metabolismo , Inflamación/metabolismo , Animales , Citocinas/metabolismo , Femenino , Factor 15 de Diferenciación de Crecimiento/genética , Humanos , Inflamación/inducido químicamente , Inflamación/genética , Leptina/metabolismo , Lipopolisacáridos/farmacología , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones TransgénicosRESUMEN
Protein ubiquitylation is a complex enzymatic process that results in the covalent attachment of ubiquitin, through Gly-76 of ubiquitin, to an varepsilonNH2 group of an internal lysine residue in a given substrate. Although E3 ligases frequently use lysines adjacent to the degron within the substrate, many substrates can be targeted to the proteasome through the polyubiquitylation of any lysine. We have assessed the role of lysine residues proximal to the cyclin D1 phosphodegron for ubiquitylation by the SCF(Fbx4/alphaB-crystallin) ubiquitin ligase and subsequent proteasome-dependent degradation of cyclin D1. The work described herein reveals a requisite role for Lys-269 (K269) for the rapid polyubiquitin-mediated degradation of cyclin D1. Mutation of Lys-269, which is proximal to the phosphodegron sequence surrounding Thr-286 in cyclin D1, not only stabilizes cyclin D1 but also triggers cyclin D1 accumulation within the nucleus, thereby promoting cell transformation. In addition, D1-K269R is resistant to genotoxic stress-induced degradation, similar to non-phosphorylatable D1-T286A, supporting the critical role for the post-translational regulation of cyclin D1 in response to DNA-damaging agents. Strikingly, although mutation of lysine 269 to arginine inhibits cyclin D1 degradation, it does not inhibit cyclin D1 ubiquitylation in vivo, showing that ubiquitylation of a specific lysine can influence substrate targeting to the 26S proteasome.