Dysregulation of amino acid metabolism upon rapid depletion of cap-binding protein eIF4E [aro10_screen]

Protein synthesis is metabolically costly, and the level of translation must match nutrient availability and cellular needs. Overall protein synthesis levels are modulated by regulating translation initiation. The cap-binding protein eIF4E—the earliest contact between mRNAs and the translation machinery—serves as one point of control, but its contributions to mRNA-specific translation regulation remain poorly understood. We acutely depleted eIF4E, which is essential in budding yeast, and observed surprisingly modest effects on cell growth and protein synthesis. Long-lived transcripts were downregulated, likely reflecting accelerated turnover, and the strongest gene-specific effects arose as secondary effects of reduced protein biosynthesis on amino acid pools. Futile cycles of amino acid synthesis and degradation were accompanied by translational activation of GCN4, which is typically induced by amino acid starvation. We further identified translational tuning of PCL5, a negative regulator of Gcn4, that provides a consistent protein-to-mRNA ratio under varying translation environments. This translational control depended in part on a uniquely long poly-(A) tract in the PCL5 5’ UTR and on poly-(A) binding protein. These results highlight the intricate interplay between translation, amino acid homeostasis, and gene regulation and uncover new layers of feedback control in cellular response to stress and nutrient availability.蛋白质合成在代谢上成本高昂，翻译水平必须与营养可获得性及细胞需求相匹配。整体蛋白质合成水平通过调控翻译起始得以调节。帽结合蛋白eIF4E（cap-binding protein eIF4E）作为信使RNA（messenger RNA，mRNA）与翻译机器间的首个结合因子，是一类调控靶点，但其在mRNA特异性翻译调控中的具体贡献仍不甚明晰。我们在出芽酵母中急性耗竭了必需基因eIF4E，意外观察到其对细胞生长与蛋白质合成的影响仅为温和效应。长寿命转录本的表达出现下调，这大概率反映了其降解速率的加速；而最显著的基因特异性效应，源于蛋白质生物合成减少对氨基酸池产生的次级效应。氨基酸合成与降解的无效循环伴随GCN4的翻译激活——GCN4通常仅在氨基酸饥饿条件下被诱导表达。我们进一步发现了PCL5的翻译调控机制：PCL5是Gcn4的负调控因子，其可在不同翻译环境下维持稳定的蛋白质-信使RNA比例。该翻译调控部分依赖于PCL5 5'非翻译区（5' untranslated region, 5' UTR）内一段独特的长多聚腺苷酸序列（poly-(A) tract），以及多聚腺苷酸结合蛋白（poly-(A) binding protein）。上述结果揭示了翻译、氨基酸稳态与基因调控间的复杂相互作用，并阐明了细胞对应激与营养可获得性产生响应的新型反馈调控层级。