Poster Presentation 44th Lorne Genome Conference 2023

Exquisite timing and mechanistic diversity of gene expression changes in response to a prototypical nutritional stress   (#234)

Yoshika Janapala 1 2 3 , Attila Horvath 1 2 , Ross D Hannan 3 4 5 6 , Eduardo Eyras 1 , Nikolay E Shirokikh 1 2 , Thomas Preiss 1 2 7
  1. Division of Genome Sciences and Cancer, The John Curtin School of Medical Research, The Australian National University, Canberra, Australian Capital Territory, Australia
  2. Shine-Dalgarno centre for RNA innovation, Australian national University, Canberra, ACT, Australia
  3. Biochemistry & Molecular Biology, Monash Biomedicine Discovery Institute, Monash University, Melbourne, Victoria, Australia
  4. John Curtin School of Medical Research, ANU, John Curtin School of Medical Research, Acton, ACT, Australia
  5. 3. Department of Biochemistry and Molecular Biology, University of Melbourne, Parkville, Victoria 3010, Australia, University of Melbourne, Melbourne, Victoria, Australia
  6. School of Biomedical Sciences, University of Queensland, St Lucia, Queensland, Australia
  7. Victor Chang Cardiac Research Institute, Sydney, NSW, Australia

The control of mRNA translation into proteins is critical for the adaptation of eukaryotic cells to environmental changes and stress conditions. Glucose starvation in yeast is one of the prototypical eukaryotic stresses. Early translation-mediated gene expression changes in glucose starvation are critically important to trigger the subsequent events leading to transcriptional reprogramming, but the mRNAs involved and the mechanisms of their selective regulation remain obscure.

In budding yeast, we analysed glucose-specific changes to transcriptome composition and mRNA translation during rapid (20 seconds) and acute starvation (600 seconds). To this end, we performed conventional RNA-seq as well as determining the distribution of diverse ribosomal complexes along mRNA by Translation Complex Profile sequencing (TCP-seq) (Janapala et al. 2021 JoVE) (Horvath, Janapala et al. 2022 bioRxiv). We used an enhanced TCP-seq protocol providing separate footprint data for singular ribosomes, and collided di-ribosomes. The method further yields separate footprints of small ribosomal subunits (SSU) in different functional states, e.g., those in polysomal complexes or attached as singletons to mRNA, or those associated with a particular translation initiation factor such as eIF4A.

The data present an in-depth collection of mRNAs involved in rapid and acute responses to glucose starvation and the enrichment of directly relevant gene ontology terms (e.g., glucose transport and synthesis, heat shock or ribosome biogenesis) in co-regulated mRNA groups. Furthermore, as our range of RNA-seq and translation complex footprint data reports on distinct functional states of mRNA, we uncover a complex picture of the modes of regulatory change and their respective timing. For example, regulation at the level of mRNA turnover occurs rapidly, whereas new mRNA synthesis, condensation into stress granules, and intervention at distinct steps of translation predominate during acute starvation.

Taken together, this work provides rich new information on the gene regulatory changes and mechanisms at play in the response to nutritional stress. Furthermore, it offers an expanded repertoire of TCP-seq approaches that should be of use in investigating gene regulation mechanisms in diverse cellular contexts.