Cross-talks between metabolic and translational controls during beige adipocyte differentiation [Ribo-seq]. Cross-talks between metabolic and translational controls during beige adipocyte differentiation [Ribo-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. Overall design: Ribo-seq to measure the translation activity 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的细胞表型与代谢特征，其具体作用机制至今尚未明确。本研究首次揭示了米色脂肪细胞中代谢调控与翻译调控之间两条此前未被发现的串扰通路。本研究通过分析前体脂肪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，发现氧化磷酸化（OXPHOS）复合物组分编码mRNA存在选择性翻译下调现象。此次下调仅局限于复合物I、III、IV与V，且与三羧酸循环（TCA cycle）相关基因的翻译增强协同发生，进而改变产热脂肪细胞线粒体中氧化磷酸化与三羧酸循环组分的化学计量比，并调控其相关代谢活性。我们通过对核糖体排布的高分辨率解析，发现谷氨酸密码子位点的核糖体暂停现象显著增强。在脂肪细胞整体分化过程中，谷氨酸消耗升高导致谷氨酸负载tRNA水平下降，进而引发核糖体暂停现象加剧。核糖体暂停会降低富含谷氨酸密码子基因的蛋白质合成效率与mRNA稳定性，例如肌动蛋白细胞骨架相关基因。实验整体设计：采用核糖体测序（Ribo-seq）检测米色脂肪细胞分化过程中的翻译活性。从腹股沟白色脂肪组织中分离脂肪前体细胞，诱导其向米色脂肪细胞分化；分别取未诱导组与分化4天、8天的样本进行文库构建，每个条件设置三次生物学重复。