Evidence of defined temporal expression patterns that lead a gram-negative cell out of dormancy

Many bacterial species are capable of forming long-lived dormant cells. The best characterized are heat and desiccation resistant spores produced by many Gram-positive species. Less characterized are dormant cysts produced by several Gram-negative species that are somewhat tolerant to increased temperature and very resistant to desiccation. While there is progress in understanding regulatory circuits that control spore germination, there is scarce information on how Gram-negative organisms emerges from dormancy. In this study, we show that R. centenum cysts germinate by emerging a pair of motile vegetative cells from a thick cyst cell wall coat similar to 6 hrs post induction of germination. Time-lapse transcriptomic analysis reveals that there is a defined temporal pattern of gene expression changes during R. centenum cyst germination. The first observable changes are increases in expression of genes for protein synthesis, an increase in expression of genes involved in the generation of a membrane potential and the use of this potential for ATP synthesis via ATPase expression. These early events are followed by expression changes that affect the cell wall and membrane composition, followed by expression changes that promote chromosome replication. Midway through germination, expression changes occur that promote the flow of carbon through the TCA cycle to generate reducing power and parallel synthesis of electron transfer components involved in oxidative phosphorylation. Finally, late expression changes promote the synthesis of a photosystem as well as flagellar and chemotaxis components for motility. Author summary Many bacteria are capable of forming long lived dormant cell types that are resistant to harsh environmental conditions. The best characterized example are spores synthesized by several Gram-positive bacteria such as Bacillus and Clostridia species. Not well characterized are long-lived desiccation resistant cysts synthesized by several Gram-negative bacteria including some pathogens. Several studies have described physiological changes that occur during the germination of Gram-positive spores, however, there are no detailed studies on physiological changes that occur during Gram-negative cyst germination. In this study, we developed a method for isolating cysts from the Gram-negative species Rhodospirillum centenum and studied genome wide changes in gene expression that occur as cysts germinate into replicative cells. This study reveals a programed set of expression changes during germination that favors an early generation of a membrane potential for energy production, coupled with expression of gene involved in cellular defense. These early events are followed by large scale physiological changes that promote cellular replication.