Abstract

AMP-activated protein kinase (AMPK) is indispensable for the development and maintenance of brown adipose tissue (BAT), and its activity is inhibited due to obesity. The isocitrate dehydrogenase 2 (IDH2) is a mitochondrial enzyme responsible for the production of α-ketoglutarate, a key intermediate metabolite integrating multiple metabolic processes. We previously found that AMPKα1 ablation reduced cellular α-ketoglutarate concentration during brown adipocyte differentiation, but the effect of AMPKα1 on Idh2 expression remains undefined. In the present study, mouse C3H10T1/2 cells were transfected with Idh2-CRISPR/Cas9, and induced to brown adipogenesis. Our data suggested that brown adipogenesis was compromised due to IDH2 deficiency in vitro, which was accompanied by down-regulation of PR-domain containing 16. Importantly, the IDH2 content was reduced in brown stromal vascular cells (BSVs) separated from AMPKα1 knockout (KO) BAT, which was associated with lower contents of histone 2B (H2B) O-GlcNAcylation and monoubiquitination. Furthermore, both GlcNAcylated-H2B (S112) and ubiquityl-histone 2B (K120) contents in the Idh2 promoter were decreased in AMPKα1 KO BSVs. Meanwhile, ectopic O-linked N-acetylglucosamine transferase (OGT) expression was positively correlated with Idh2 expression, while OGT (T444A) mutation abolished the regulatory effect of AMPKα1 on Idh2. In vivo, reduced AMPKα1 activity and lower IDH2 abundance were observed in BAT of obese mice when compared with those in control mice. Taken together, our data demonstrated that IDH2 is necessary for brown adipogenesis and that AMPKα1 deficiency attenuates Idh2 expression, which might be by suppressing H2B O-GlcNAcylation modification.

中文翻译：

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AMPKα1在褐色脂肪形成过程中通过H2B O-GlcNAcylation调节Idh2转录

摘要

AMP激活的蛋白激酶（AMPK）对于棕色脂肪组织（BAT）的发育和维持是必不可少的，并且其活性由于肥胖而受到抑制。异柠檬酸脱氢酶2（IDH2）是负责产生α-酮戊二酸的线粒体酶，α-酮戊二酸是整合多个代谢过程的关键中间代谢物。我们先前发现AMPKα1消融可降低棕色脂肪细胞分化过程中细胞α-酮戊二酸的浓度，但AMPKα1对Idh2表达的影响尚不确定。在本研究中，用Idh2- CRISPR / Cas9转染小鼠C3H10T1 / 2细胞，并诱导其成褐色。我们的数据表明，由于体外IDH2缺乏，棕色脂肪形成受到损害。，伴随着含有16的PR结构域的下调。重要的是，从AMPKα1敲除（KO）BAT分离的棕色基质血管细胞（BSV）中IDH2含量降低，这与组蛋白2B（H2B）的含量降低有关）O-GlcNAcylation和单泛素化。此外，在AMPKα1KO BSV中，Idh2启动子中的GlcNAcylated-H2B（S112）和泛素化组蛋白2B（K120）含量均降低。同时，异位O-连接的N-乙酰氨基葡萄糖转移酶（OGT）表达与Idh2表达正相关，而OGT（T444A）突变消除了AMPKα1对Idh2的调节作用。体内与对照组相比，肥胖小鼠的BAT中AMPKα1活性降低，IDH2丰度降低。两者合计，我们的数据表明IDH2是棕色脂肪形成所必需的，而AMPKα1缺乏会减弱Idh2表达，这可能是通过抑制H2B O-GlcNAcylation修饰而实现的。