Regulation of Adipocyte Differentiation by Activation of Serotonin (5-HT) Receptors 5-HT2AR and 5-HT2CR and Involvement of MicroRNA-448-Mediated Repression of KLF5
Regulation of Adipocyte Differentiation by Activation of Serotonin (5-HT) Receptors 5-HT2AR and 5-HT2CR and Involvement of MicroRNA-448-Mediated Repression of KLF5
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DOI：
10.1210/me.2010-0054
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发表时间：
2010-10-01
影响因子：
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通讯作者：
Kimura, Takeshi
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文献类型：
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作者：
Kinoshita, Minako;Ono, Koh;Kimura, Takeshi
将逆转录病毒插入介导的随机诱变应用于 3T3-L1 前脂肪细胞，以更好地了解肥胖的分子基础（单个脂肪细胞的扩张）。我们发现色氨酸羟化酶-1（色氨酸羟化酶-1）是一种合成血清素（5-HT）的限速酶，在脂肪细胞中表达，并且是脂肪细胞分化所必需的。 5-HT 2A 型受体 (5-HT2AR) 拮抗剂酮色林和 5-HT2cR 拮抗剂 SB-242084 可抑制脂肪细胞分化。由于5-HT2cR mRNA水平在脂肪细胞分化过程中上调，并且微小RNA（miR）-448位于Htr2c的第四个内含子中，因此我们还研究了miR-448在3T3-L1细胞中的作用。通过生物信息学方法，Kruppel 样因子 5 (KLF5) 被确定为 miR-448 的潜在靶标。使用荧光素酶报告基因测定，我们证实 miR-448 靶向 Klf5 3'-翻译区。 miR-448的过表达降低了Klf5的表达和脂肪细胞分化，这通过脂肪形成基因的表达减少和甘油三酯积累得到证实。为了检查 miR-448 功能的丧失，我们构建了一个诱饵基因，该诱饵基因在巨细胞病毒启动子的控制下，在荧光素酶基因的 3'-非翻译区中具有 miR-448 的串联互补序列。当将 miR-448 诱饵基因引入 3T3-L1 前脂肪细胞时，KLF5 上调，甘油三酯浓度增加。在这项研究中，我们确定了 5-HT、5-HT2AR 和 5-HT2CR 对脂肪细胞分化的调节。 miR-448 介导的 KLF5 抑制被确定为脂肪细胞分化的负调节因子。 （分子内分泌学24：1978-1987，2010）
Retrovirus insertion-mediated random mutagenesis was applied in 3T3-L1 preadipocyte cells to better understand the molecular basis of obesity (the expansion of individual adipocytes). We found that tryptophan hydroxylase-1, a rate-limiting enzyme for the synthesis of serotonin (5-HT), is expressed in adipocytes and is required for their differentiation. A 5-HT type 2A receptor (5-HT2AR) antagonist, ketanserin, and a 5-HT2cR antagonist, SB-242084, inhibited adipocyte differentiation. Because 5-HT2cR mRNA levels are up-regulated during adipocyte differentiation and micro-RNA (miR)-448 is located in the fourth intron of Htr2c, we also studied the role of miR-448 in 3T3-L1 cells. Through a bioinformatics approach, Kruppel-like factor 5 (KLF5) was identified as a potential target of miR-448. Using a luciferase reporter assay, we confirmed that miR-448 targets the Klf5 3'-intranslated region. Overexpression of miR-448 reduced the expression of Klf5 and adipocyte differentiation, which was confirmed by the reduced expression of adipogenic genes and triglyceride accumulation. To examine the loss of miR-448 function, we constructed a decoy gene that had tandem complementary sequences for miR-448 in the 3'-untranslated region of a luciferase gene under the control of a cytomegalovirus promoter. When the miR-448 decoy gene was introduced into 3T3-L1 preadipocytes, KLF5 was up-regulated and triglyceride concentration was increased. In this study, we identified the regulation of adipocyte differentiation by 5-HT, 5-HT2AR, and 5-HT2CR. miR-448-mediated repression of KLF5 was identified as a negative regulator for adipocyte differentiation. (Molecular Endocrinology 24: 1978-1987, 2010)