Sustained ppGpp production underpins months-long survival of a bacterium in growth arrest

When growing bacteria start to reach stationary phase, the nucleotide guanosine tetra-phosphate (ppGpp) accumulates intracellularly and regulates the transition of cells from growth to growth arrest. Because commonly studied bacteria remain viable in stationary phase only briefly under laboratory conditions, the role of ppGpp in sustaining long-term bacterial survival after growth arrest has not been widely studied. Rhodopseudomonas palustris strain CGA009 is a phototrophic alpha-proteobacterium that survives under anaerobic conditions for months when not growing due to carbon starvation if provided light. When we quantified intracellular ppGpp in growing and growth-arrested R. palustris, we found that it was undetectable in growing cells but accumulated to about 100 µM when cells ran out of carbon and entered the stationary phase. These elevated levels of ppGpp were maintained over a 60-day period of growth arrest. Intracellular GTP was 100-200 µM in growth-arrested cells, and ATP was at 2-4 mM. ppGpp had global effects on gene expression, with over half of the genes in the R. palustris genome being activated or repressed by ppGpp in stationary phase cells. These results suggest that, in addition to its known role in facilitating the transition of bacteria from growth to stationary phase and accompanying stress responses, ppGpp is important for prolonging bacterial survival in stationary phase.

The molecular basis for long-term survival of starved bacteria is not well understood. The bacterium Rhodopseudomonas palustris has a metabolism in which it survives in growth arrest for months, as long as it can generate ATP by photophosphorylation. This allowed us to determine if the nucleotide ppGpp, produced by bacteria in response to nutrient starvation, plays a role in supporting the viability of non-growing bacteria for long periods. We found that ppGpp accumulated as R. palustris entered growth arrest, and its intracellular levels were maintained for 60 days. ppGpp regulated the expression of over half the genes in the R. palustris genome in stationary phase cells. This work expands our concept of the effects of ppGpp on bacterial physiology to encompass an important role in long-term bacterial starvation-survival and longevity.