Enhanced uptake of potassium or glycine betaine or export of cyclic-di-AMP restores osmoresistance in a high cyclic-di-AMP Lactococcus lactis mutant

• Main Authors: Pham, Huong Thi, Nhiep, Nguyen Thi Hanh, Vu, Thu Ngoc Minh, Huynh, TuAnh Ngoc, Zhu, Yan, Huynh, Anh Le Diep, Chakrabortti, Alolika, Marcellin, Esteban, Lo, Raquel, Howard, Christopher B., Bansal, Nidhi, Liang, Zhao-Xun, Turner, Mark S., Woodward, Joshua J.
• Other Authors: Garsin, Danielle A.
• Format: Article
• Language: English
• Published: 2018
• Subjects: Lactococcus Mutant; Osmoresistance; DRNTU::Science::Biological sciences
• Online Access: https://hdl.handle.net/10356/89801; http://hdl.handle.net/10220/46391
• Institution: Nanyang Technological University

The broadly conserved bacterial signalling molecule cyclic-di-adenosine monophosphate (c-di-AMP) controls osmoresistance via its regulation of potassium (K+) and compatible solute uptake. High levels of c-di-AMP resulting from inactivation of c-di-AMP phosphodiesterase activity leads to poor growth of bacteria under high osmotic conditions. To better understand how bacteria can adjust in response to excessive c-di-AMP levels and to identify signals that feed into the c-di-AMP network, we characterised genes identified in a screen for osmoresistant suppressor mutants of the high c-di-AMP Lactococcus ΔgdpP strain. Mutations were identified which increased the uptake of osmoprotectants, including gain-of-function mutations in a Kup family K+ importer (KupB) and inactivation of the glycine betaine transporter transcriptional repressor BusR. The KupB mutations increased the intracellular K+ level while BusR inactivation increased the glycine betaine level. In addition, BusR was found to directly bind c-di-AMP and repress expression of the glycine betaine transporter in response to elevated c-di-AMP. Interestingly, overactive KupB activity or loss of BusR triggered c-di-AMP accumulation, suggesting turgor pressure changes act as a signal for this second messenger. In another group of suppressors, overexpression of an operon encoding an EmrB family multidrug resistance protein allowed cells to lower their intracellular level of c-di-AMP through active export. Lastly evidence is provided that c-di-AMP levels in several bacteria are rapidly responsive to environmental osmolarity changes. Taken together, this work provides evidence for a model in which high c-di-AMP containing cells are dehydrated due to lower K+ and compatible solute levels and that this osmoregulation system is able to sense and respond to cellular water stress.