Queuosine biosynthetic enzyme, QueE moonlights as a cell division regulator

Abstract

In many organisms, stress responses to adverse environments can trigger secondary functions of certain proteins by altering protein levels, localization, activity, or interaction partners. Escherichia coli cells respond to the presence of specific cationic antimicrobial peptides by strongly activating the PhoQ/PhoP two-component signaling system, which regulates genes important for growth under this stress. As part of this pathway, a biosynthetic enzyme called QueE, which catalyzes a step in the formation of queuosine (Q) tRNA modification is upregulated. When cellular QueE levels are high, it co-localizes with the central cell division protein FtsZ at the septal site, blocking division and resulting in filamentous growth. Here we show that QueE affects cell size in a dose-dependent manner. Using alanine scanning mutagenesis of amino acids in the catalytic active site, we pinpoint particular residues in QueE that contribute distinctly to each of its functions - Q biosynthesis or regulation of cell division, establishing QueE as a moonlighting protein. We further show that QueE orthologs from enterobacteria like Salmonella typhimurium and Klebsiella pneumoniae also cause filamentation in these organisms, but the more distant counterparts from Pseudomonas aeruginosa and Bacillus subtilis lack this ability. By comparative analysis of E. coli QueE with distant orthologs, we elucidate a unique region in this protein that is responsible for QueEs secondary function as a cell division regulator. A dual-function protein like QueE is an exception to the conventional model of "one gene, one enzyme, one function", which has divergent roles across a range of fundamental cellular processes including RNA modification and translation to cell division and stress response. Author SummaryIn stressful environments, proteins in many organisms can take on extra roles. When Escherichia coli bacteria are exposed to antimicrobial compounds, the cell activates the PhoQ/PhoP signaling system, increasing the production of an enzyme called QueE. QueE is usually involved in the formation of queuosine (Q) tRNA modification. When cells make abundant QueE, it interacts with a vital division protein, FtsZ, disrupting division and causing elongation - a "moonlighting" function. Detailed study of QueE reveals specific regions involved in Q biosynthesis or cell division. QueE in organisms closely related to E. coli also has dual roles, while distant relatives are unifunctional. Comparative analysis identifies a unique E. coli QueE region regulating cell division. This study shows QueEs versatility in linking and impacting distinct cellular processes such as RNA metabolism, protein translation, cell division, and stress response.