Having multiple rounds of translation of the same mRNA is a key feature of the Central Dogma that creates substantial dynamic complexities along with opportunities for regulation related to ribosome pausing and stalling at specific sequences. The molecular mechanisms controlling these critical processes and the principles guiding their evolution remain among the least understood facets of cellular physiology. We herein use a combination of genetic, genomic, physiological, and biochemical methods to show that regulation of ribosome pausing at specific amino acid sequences can produce ~2-fold changes in protein expression levels that have a strong influence on cell growth and therefore evolutionary fitness. We demonstrate, both in vivo and in vitro, that the ABCF protein EttA directly controls the translation of mRNAs coding for a subset of enzymes in the tricarboxylic acid (TCA) cycle and its glyoxylate shunt, which dramatically modulates growth in some chemical environments. It also modulates expression of specific proteins involved in metabolically related physiological and stress-response pathways. These regulatory activities are mediated by EttA rescuing ribosomes paused at specific patterns of negatively charged residues within the first 30 amino acids of nascent proteins. The previously established dependency of EttA9s activity on ADP:ATP ratio can modulate these effects based on cellular energy status. We thus establish a new global regulatory paradigm based on sequence-specific modulation of translational pausing.

Global regulation via modulation of ribosome pausing by EttA