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. Overall design: To determine the impact of reduced eIF4E levels on ribosome occupancy transcriptome-wide, we established a eIF4E-AID (auxin inducible degron) yeast strain. We then depleted eIF4E by adding IAA for either 1 or 8 hours. We determined changes in translation and RNA abundance by ribosome profiling and RNA-seq, respectively. A DMSO treated sample was used as a control and both conditions (IAA and DMSO) were performed in duplicate.