Center for Molecular Medicine, Maine Medical Center Research Institute, Scarborough, Maine.
The Graduate School of Biomedical Science and Engineering, University of Maine, Orono, Maine.
J Cell Biochem. 2018 Mar;119(3):2636-2645. doi: 10.1002/jcb.26429. Epub 2017 Dec 4.
Highly variable expression of mesoderm-specific transcript (Mest) in adipose tissue among genetically homogeneous mice fed an obesogenic diet, and its positive association with fat mass expansion, suggests that Mest is an epigenetic determinant for the development of obesity. Although the mechanisms by which MEST augments fat accumulation in adipocytes have not been elucidated, it has sequence homology and catalytic peptide motifs which suggests that it functions as an epoxide hydrolase or as a glycerol- or acylglycerol-3-phosphate acyltransferase. To better understand MEST function, detailed studies were performed to precisely define the intracellular organelle localization of MEST using immunofluorescence confocal microscopy. Lentiviral-mediated expression of a C-terminus Myc-DDK-tagged MEST fusion protein expressed in 3T3-L1 preadipocytes/adipocytes, and ear-derived mesenchymal stem cells (EMSC) from mice was observed in the endoplasmic reticulum (ER) membranes and is consistent with previous studies showing endogenous MEST in the membrane fraction of adipose tissue. MEST was not associated with the Golgi apparatus or mitochondria; however, frequent contacts were observed between MEST-positive ER and mitochondria. MEST-positive domains were also shown on the plasma membrane (PM) of non-permeabilized cells but they did not co-localize with ER-PM bridges. Post-adipogenic differentiated 3T3-L1 adipocytes and EMSC showed significant co-localization of MEST with the lipid droplet surface marker perilipin at contact points between the ER and lipid droplet. Identification of MEST as an ER-specific protein that co-localizes with lipid droplets in cells undergoing adipogenic differentiation supports a function for MEST in the facilitation of lipid accumulation and storage in adipocytes.
高脂肪饮食喂养的遗传同质性肥胖小鼠的脂肪组织中中胚层特异性转录物（Mest）的表达高度可变，并且与脂肪质量扩张呈正相关，这表明 Mest 是肥胖发生的表观遗传决定因素。虽然尚未阐明 MEST 增强脂肪细胞脂肪堆积的机制，但它具有序列同源性和催化肽基序，这表明它作为环氧化物水解酶或甘油酯或酰基甘油-3-磷酸酰基转移酶发挥作用。为了更好地了解 MEST 的功能，进行了详细的研究，使用免疫荧光共焦显微镜精确定义 MEST 的细胞内细胞器定位。用慢病毒介导表达的 3T3-L1 前脂肪细胞/脂肪细胞中的 C 端 Myc-DDK 标记的 MEST 融合蛋白，以及从小鼠耳朵衍生的间充质干细胞（EMSC）进行观察，发现其定位于内质网（ER）膜，这与先前的研究一致，表明脂肪组织中存在内源性 MEST。MEST 与高尔基体或线粒体无关；然而，在 ER 阳性和线粒体之间观察到频繁的接触。在未通透细胞的质膜（PM）上也观察到 MEST 阳性结构域，但它们与 ER-PM 桥不共定位。脂肪生成分化后的 3T3-L1 脂肪细胞和 EMSC 显示 MEST 与 ER 和脂质滴之间接触点处的脂质滴表面标记蛋白 perilipin 显著共定位。鉴定 MEST 作为一种 ER 特异性蛋白，在经历脂肪生成分化的细胞中与脂质滴共定位，支持 MEST 在促进脂肪细胞中脂质积累和储存的功能。
