SIRT1 Deacetylates and Inhibits SREBP-1C Activity in Regulation of Hepatic Lipid Metabolism

The SIRT1 deacetylase inhibits fat synthesis and stimulates fat oxidation in response to fasting, but the underlying mechanisms remain unclear. Here we report that SREBP-1c, a key lipogenic activator, is an in vivo target of SIRT1. SIRT1 interaction with SREBP-1c was increased during fasting and decreased upon feeding, and consistently, SREBP-1c acetylation levels were decreased during fasting in mouse liver. Acetylated SREBP-1c levels were also increased in HepG2 cells treated with insulin and glucose to mimic feeding conditions, and down-regulation of p300 by siRNA decreased the acetylation. Depletion of hepatic SIRT1 by adenoviral siRNA increased acetylation of SREBP-1c with increased lipogenic gene expression. Tandem mass spectrometry and mutagenesis studies revealed that SREBP-1c is acetylated by p300 at Lys-289 and Lys-309. Mechanistic studies using acetylation-defective mutants showed that SIRT1 deacetylates and inhibits SREBP-1c transactivation by decreasing its stability and its occupancy at the lipogenic genes. Remarkably, SREBP-1c acetylation levels were elevated in diet-induced obese mice, and hepatic overexpression of SIRT1 or treatment with resveratrol, a SIRT1 activator, daily for 1 week decreased acetylated SREBP-1c levels with beneficial functional outcomes. These results demonstrate an intriguing connection between elevated SREBP-1c acetylation and increased lipogenic gene expression, suggesting that abnormally elevated SREBP-1c acetylation increases SREBP-1c lipogenic activity in obese mice. Reducing acetylation of SREBP-1c by targeting SIRT1 may be useful for treating metabolic disorders, including fatty liver, obesity, and type II diabetes.     

RNA特异腺苷脱氨酶1(p150亚型)抑制肝细胞脂质合成

目的:在油酸诱导的肝细胞脂肪变模型中,检测RNA特异腺苷脱氨酶1 p150亚型(ADAR1-p150)高表达细胞系中脂肪合成的变化。方法:利用本课题组前期摸索的油酸刺激人胚胎肝细胞L-02细胞系脂肪变的条件,q RT-PCR和Western-Blot检测油酸刺激组和对照组ADAR1-p150表达变化;将构建成功的ADAR1-p150过表达慢病毒载体GV166-ADAR1-p150及空载体病毒GV166-control感染L-02细胞,检测感染细胞中ADAR1-p150的m RNA和蛋白表达水平;通过油红O染色和BODIPY染色观察L-02 ADAR1-p150和L-02 control细胞中脂滴形成,并进一步利用高内涵系统检测其荧光强度,对脂滴合成作定量分析。结果:L-02细胞在油酸刺激后ADAR1-p150的m RNA和蛋白水平降低;成功构建ADAR1-p150过表达慢病毒载体GV166-ADAR1-p150及空载体病毒GV166-control,q RT-PCR及Western-Blot检测显示病毒转染GV166-ADAR1-p150后ADAR1-p150在细胞中的表达水平显著升高;油红O染色和BODIPY染色发现L-02 ADAR1-p150较L-02 control细胞胞质中脂滴数量减少。高内涵筛选系统检测提示L-02 ADAR1-p150组中脂滴的荧光强度明显较L-02 control组低。结论:成功构建ADAR1-p150过表达稳定转染L-02细胞系,并证实高表达ADAR1-p150能够抑制脂肪合成。     

FABP5 is a critical regulator of methionine- and estrogen-induced SREBP-1c gene expression in bovine mammary epithelial cells

The intracellular fatty acid-binding proteins (FABPs) are a well-conserved family that function as lipid chaperones. Ongoing studies are focused on identification of the mechanistic complexity and vast biological diversity of different isoforms of FABPs. However, the molecular mechanism of FABP5 in the regulation of milk fat synthesis in the mammary gland of dairy cows is still largely unknown. Here, we report that FABP5 acts as a critical regulator of terol response element-binding protein-1c (SREBP-1c) gene expression induced by methionine (Met) and estrogen (E2) in bovine mammary epithelial cells (BMECs). We observed that the expression of FABP5 was markedly higher in dairy cow mammary tissue during the lactating period than the puberty period and the dry period. FABP5 is located in the cytoplasm, and Met and E2 significantly increase the protein levels of FABP5 in BMECs. Using gene function study approaches, we revealed that FABP5 positively regulates SREBP-1c gene expression and promotes milk fat synthesis. We confirmed that FABP5 is required for Met- and E2-induced SREBP-1c gene expression and milk fat synthesis. We further uncovered that fatty acids are needed for FABP5-mediated SREBP-1c gene expression. Thus, our study demonstrates that FABP5 is a critical regulator of Met- and E2-induced SREBP-1c gene expression leading to milk fat synthesis.