A novel bacterial cell death pathway targeted by a poisoned cell cycle sentinel

The relentless increase in bacterial resistance is a major global health threat. The expanding number of people that succumb to bacterial infections due to a lack of effective treatment is extremely alarming. Mankind is therefore in urgent need of novel and innovative antibacterial compounds. The development of such antibacterial therapies could be aided by a deepened understanding of bacterial physiology. Besides an obvious fundamental interest, increased insight into how bacteria proliferate and sustain viability could reveal new points of interference that can be targeted by future antibiotics. While it has been firmly established that the GTPase Obg is vital for bacterial proliferation and plays an important role in the bacterial cell cycle, its precise cellular function is currently unknown. Nonetheless, efforts are being made to identify antibacterial compounds that target the Obg protein. Investigation of Obg functionality is hampered by the inability to create obg deletion mutants, the fact that depletion and overexpression of Obg often have similar effects and the occurrence of very pleiotropic phenotypes. We here use a dominant-negative isoform of the ObgE protein (Obg of Escherichia coli), ObgE*, as a tool to study ObgE function. Insights gleaned from this approach could support the development of Obg-targeting therapies. The mutation present in ObgE* is intriguing because it changes ObgE from a protein that is essential for bacterial survival to a highly lethal one. Upon expression of ObgE* in E. coli, cells rapidly lose viability because ObgE* irreversibly blocks progression through the cell cycle. It most likely does so by disturbing the assembly of FtsZ at midcell, thereby inhibiting cell division. Since the mutant ObgE protein described here strongly interferes with normal cell cycle progression, and because ObgE is known to be involved in cell cycle control, we hypothesize that the normal role of wild-type ObgE in the bacterial cell cycle is disturbed by the mutation present in ObgE*. Studying how ObgE* affects cell cycle progression and cell division can thus provide insight into the enigmatic role of ObgE in these processes. ObgE*-mediated toxicity can be almost completely neutralized by mutations in lpxA. The LpxA gene product catalyzes a reversible reaction that, in the forward direction, uses peptidoglycan and phospholipid precursors to produce an intermediate in lipopolysaccharide synthesis. LpxA therefore functions as a hub in cell envelope metabolism and changes in LpxA function could affect not only lipopolysaccharide synthesis but also the production of peptidoglycan and/or membrane phospholipids. Since changes in cell envelope metabolism restore the cell cycle progression defect imposed by ObgE*, our results strongly implicate cell envelope homeostasis in regulation of the bacterial cell cycle. This possible connection has been severely understudied but is potentially a very significant factor in bacterial proliferation. In conclusion, our results reveal that ObgE* inhibits cell cycle progression at the stage of cell division most likely by interfering with cell envelope metabolism. A connection between ObgE and the bacterial cell envelope has never been reported before. Moreover, although it is known that ObgE influences the cell cycle, this is the first time that Obg is suggested to affect cell division directly. These newly discovered connections are therefore of high interest for further research on the Obg protein and will likewise increase our understanding of the bacterial cell cycle and, more specifically, how it is influenced by the cell envelope. Importantly, Obg, the bacterial cell cycle, cell division and the cell envelope are all considered highly attractive targets for antibiotic development. Uncovering previously unknown connections between them could therefore lead to the discovery of new points of interference that can be targeted by novel antibacterial compounds.nrpages: 236status: publishe