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AGO HITS-CLIP reveals distinct miRNA regulation of white and brown adipose tissue identity
Genes & Development ( IF 7.5 ) Pub Date : 2021-05-01 , DOI: 10.1101/gad.345447.120
Sean O'Connor 1 , Elisabeth A Murphy 2 , Sarah K Szwed 1, 3 , Matt Kanke 4 , François Marchildon 1 , Praveen Sethupathy 4 , Robert B Darnell 2 , Paul Cohen 1
Affiliation  

MicroRNAs (miRNAs) are short, noncoding RNAs that associate with Argonaute (AGO) to influence mRNA stability and translation, thereby regulating cellular determination and phenotype. While several individual miRNAs have been shown to control adipocyte function, including energy storage in white fat and energy dissipation in brown fat, a comprehensive analysis of miRNA activity in these tissues has not been performed. We used high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP) to comprehensively characterize the network of high-confidence, in vivo mRNA:miRNA interactions across white and brown fat, revealing >20,000 unique AGO binding sites. When coupled with miRNA and mRNA sequencing, we found an inverse correlation between depot-enriched miRNAs and their targets. To illustrate the functionality of our HITS-CLIP data set in identifying specific miRNA:mRNA interactions, we show that miR-29 is a novel regulator of leptin, an adipocyte-derived hormone that coordinates food intake and energy homeostasis. Two independent miR-29 binding sites in the leptin 3′ UTR were validated using luciferase assays, and miR-29 gain and loss of function modulated leptin mRNA and protein secretion in primary adipocytes. This work represents the only experimentally generated miRNA targetome in adipose tissue and identifies multiple regulatory pathways that may specify the unique identities of white and brown fat.

中文翻译:

AGO HITS-CLIP 揭示了白色和棕色脂肪组织特性的不同 miRNA 调控

MicroRNA (miRNA) 是与 Argonaute (AGO) 相关的短非编码 RNA,可影响 mRNA 的稳定性和翻译,从而调节细胞测定和表型。虽然已显示几种单独的 miRNA 控制脂肪细胞功能,包括白色脂肪中的能量储存和棕色脂肪中的能量耗散,但尚未对这些组织中的 miRNA 活性进行综合分析。我们使用通过交联免疫沉淀 (HITS-CLIP) 分离的 RNA 的高通量测序来全面表征高可信度的网络,体内 mRNA:miRNA 在白色和棕色脂肪中的相互作用,揭示了 >20,000 个独特的 AGO 结合位点。当与 miRNA 和 mRNA 测序相结合时,我们发现富含仓库的 miRNA 与其靶标之间呈负相关。为了说明我们的 HITS-CLIP 数据集在识别特定 miRNA:mRNA 相互作用方面的功能,我们表明 miR-29 是瘦素的新型调节剂,瘦素是一种脂肪细胞衍生的激素,可协调食物摄入和能量稳态。使用荧光素酶测定法验证了瘦素 3' UTR 中两个独立的 miR-29 结合位点,并且 miR-29 功能的获得和丧失调节了初级脂肪细胞中的瘦素 mRNA 和蛋白质分泌。这项工作代表了脂肪组织中唯一通过实验生成的 miRNA 靶向组,并确定了多种调控途径,可以指定白色和棕色脂肪的独特身份。使用荧光素酶测定法验证了瘦素 3' UTR 中两个独立的 miR-29 结合位点,并且 miR-29 功能的获得和丧失调节了初级脂肪细胞中的瘦素 mRNA 和蛋白质分泌。这项工作代表了脂肪组织中唯一通过实验生成的 miRNA 靶向组,并确定了多种调控途径,可以指定白色和棕色脂肪的独特身份。使用荧光素酶测定法验证了瘦素 3' UTR 中两个独立的 miR-29 结合位点,并且 miR-29 功能的获得和丧失调节了初级脂肪细胞中的瘦素 mRNA 和蛋白质分泌。这项工作代表了脂肪组织中唯一通过实验生成的 miRNA 靶向组,并确定了多种调控途径,可以指定白色和棕色脂肪的独特身份。
更新日期:2021-05-03
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