Regulation of mRNA translation is crucial for eukaryotic cells adapting to environmental changes like glucose starvation. Glucose scarcity triggers significant changes in mRNA translation, stability, and transcription in budding yeast. However, the regulatory mechanisms and selective control of mRNAs during the initial (20 seconds) and acute (600 seconds) phases of starvation are not well understood.
We performed RNA-seq on Total Cell Lysate (TCL) and Clarified Cell Lysate (CCL) to track mRNA abundance, providing insights into mRNA turnover, new transcription, and sequestration into stress granules. The Translated Pool (TP) of mRNAs actively engaged in translation was analysed using an enhanced Translation Complex Profiling sequencing (eTCP-seq) technique. The eTCP-seq data were analysed with a machine learning approach, yielding a metric for mRNA translation output termed Stochastic Translation Efficiency (STE)1. Polysome profiling, nanopore direct RNA sequencing, and RT-qPCR corroborated our findings.
Our RNA-seq data highlights immediate selective mRNA stability control upon starvation. Contrary to expectations of a translational shutdown, mRNAs related to ribosomal protein and ribosome biogenesis (RiBi) were preserved as early as 20 seconds, with pronounced increases at 600 seconds. Our analysis delineates mRNA shifts from the general context (TCL) to the cytoplasmic setting (CCL), pinpointing mRNAs selectively retained or excluded in CCL. It reveals a strategic use of mRNA condensation to prioritize protein synthesis initially, then shifting to sugar transport for energy uptake. STE differentiated translational adjustments from other mRNA alterations. At 20 seconds starvation, RiBi and stress granule-related mRNAs were up-regulated, while endoplasmic reticulum (ER), storage vacuole, and metabolic process mRNAs were downregulated. At 600 seconds, RiBi mRNAs were downregulated while oxidative metabolism and ER membrane categories were upregulated, balancing immediate survival mechanisms with resource conservation.
Our findings uncover a multifaceted and intricate interplay between mRNA transcription, turnover and translation to preserve resources for optimal adaptation to starvation.