Table_1_Bacillus subtilis forms twisted cells with cell wall integrity defects upon removal of the molecular chaperones DnaK and trigger factor.DOCX

The protein homeostasis network ensures a proper balance between synthesis, folding, and degradation of all cellular proteins. DnaK and trigger factor (TF) are ubiquitous bacterial molecular chaperones that assist in protein folding, as well as preventing protein misfolding and aggregation. In Escherichia coli, DnaK and TF possess partially overlapping functions. Their combined depletion results in proteostasis collapse and is synthetically lethal at temperatures above 30°C. To increase our understanding on how proteostasis is maintained in Gram-positive bacteria, we have investigated the physiological effects of deleting dnaK and tig (encoding for DnaK and TF) in Bacillus subtilis. We show that combined deletion of dnaK and tig in B. subtilis is non-lethal, but causes a severe pleiotropic phenotype, including an aberrant twisted and filamentous cell morphology, as well as decreased tolerance to heat and to cell wall active antibiotics and hydrolytic enzymes, indicative of defects in cell wall integrity. In addition, cells lacking DnaK and TF have a much smaller colony size due to defects in motility. Despite these physiological changes, we observed no major compromises in important cellular processes such as cell growth, FtsZ localization and division and only moderate defects in spore formation. Finally, through suppressor analyses, we found that the wild-type cell shape can be partially restored by mutations in genes involved in metabolism or in other diverse cellular processes.