Cross-talks between metabolic and translational controls during beige adipocyte differentiation [RNA-seq]

Whether and how regulatory events at the translation stage shape the cellular and metabolic features of thermogenic adipocytes is hardly understood. In this study, we report two hitherto unidentified cross-talk pathways between metabolic and translation regulation in beige adipocytes. By analysing temporal profiles of translation activity and protein level changes during precursor-to-beige differentiation, we found selective translational down-regulation of OXPHOS component-coding mRNAs. The down-regulation restricted to Complexes I, III, IV, and V, is coordinated with enhanced translation of TCA cycle genes, engendering distinct stoichiometry of OXPHOS and TCA cycle components and altering the related metabolic activities in mitochondria of thermogenic adipocytes. Our high-resolution description of ribosome positioning unveiled potentiated ribosome pausing at glutamate codons. The increased stalling is driven by elevated glutamate consumption that diminishes glutamate-charged tRNA during pan-adipocyte differentiation. The ribosome pauses decrease protein synthesis and mRNA stability of glutamate codon-rich genes, such as actin cytoskeleton-associated genes. RNA-seq of total RNA to measure the RNA levels upon beige adipocyte differentiation. Adipocyte precursor cells were isolated from inguinal white adipose tissue, and induction of beige adipocyte differentiation was performed. Non-induced and differentiated samples for 4 or 8 days were used for library preparation. For each condition, triplicates were constructed.

翻译阶段的调控事件是否以及如何塑造产热脂肪细胞（thermogenic adipocytes）的细胞与代谢特征，目前仍鲜有研究阐明。本研究报道了米色脂肪细胞中代谢调控与翻译调控之间两条迄今尚未被发现的交叉对话通路。通过分析脂肪前体细胞向米色脂肪细胞分化过程中翻译活性与蛋白质水平变化的时序特征，我们发现编码氧化磷酸化（OXPHOS）复合物组分的mRNA存在选择性翻译下调现象。该下调仅局限于复合物I、III、IV和V，且与三羧酸循环（TCA）相关基因的翻译增强协同发生，由此形成氧化磷酸化与三羧酸循环组分的独特化学计量比，并改变产热脂肪细胞线粒体中的相关代谢活性。我们通过对核糖体定位的高分辨率分析，揭示了谷氨酸密码子处的核糖体暂停现象显著增强。这种暂停现象的增强，源于脂肪细胞分化全过程中谷氨酸消耗量升高，导致谷氨酸负载型tRNA水平降低。核糖体暂停会降低富含谷氨酸密码子的基因（如肌动蛋白细胞骨架相关基因）的蛋白质合成速率与mRNA稳定性。本研究通过总RNA测序（RNA-seq）检测米色脂肪细胞分化过程中的RNA水平变化：实验从腹股沟白色脂肪组织中分离得到脂肪前体细胞，并诱导其向米色脂肪细胞分化；未诱导样本以及诱导分化4天、8天的样本均用于文库构建，每个实验条件均设置3次生物学重复。