Recent studies have revealed that several Gram-negative species utilize variations of the well-known chemotaxis signaling cascade to switch lifestyles in order to survive environmental stress. Two survival strategies covered in this review are the development of dormant cyst cells and biofilm formation. Both of these two types of structures involve exopolysaccharide-mediated cell-cell interactions which result in multicellular communities that confer resistance to stress conditions such as desiccation and antibiotics. This review is centered on recent advances in the understanding of phosphate flow and novel output signals in chemosensory signaling pathways that are involved in cyst formation and biofilms.

Genomic and genetic analyses have demonstrated that many species contain multiple chemotaxis-like signal transduction cascades that likely control processes other than chemotaxis. The Che3 signal transduction cascade from Rhodospirillum centenum is one such example that regulates development of dormant cysts. This Che-like cascade contains two hybrid response regulator-histidine kinases, CheA3 and CheS3, and a single-domain response regulator CheY3. We demonstrate that cheS3 is epistatic to cheA3 and that only CheS3∼P can phosphorylate CheY3. We further show that CheA3 derepresses cyst formation by phosphorylating a CheS3 receiver domain. These results demonstrate that the flow of phosphate as defined by the paradigm E. coli chemotaxis cascade does not necessarily hold true for non-chemotactic Che-like signal transduction cascades.

Rhodospirillum centenum forms metabolically dormant cysts under unfavorable growth conditions such as desiccation or nutrient starvation. The development of cysts is tightly regulated and involves a cyst-repressing chemotaxis-like signal transduction pathway called the Che3 signaling cascade. The Che3 cascade is comprised of a methyl chemoreceptor (MCP3), receptor-methylating/demethylating proteins CheB3 and CheR3, two CheW3 linker proteins, a CheA3-CheY hybrid histidine kinase, and a single-domain response regulator, CheY3. In addition to Che-like components, the Che3 cascade also contains a second hybrid histidine kinase, CheS3. Recent biochemical and genetic studies show that CheA3 does not serve as a phosphor donor for CheY3; instead, CheA3 inhibits a CheS3→CheY3 two-component system by phosphorylating an inhibitory receiver domain of CheS3. In this study, we show that in addition to phosphorylation by CheA3, the phosphorylation state of CheS3 is also regulated by the cellular energy level as quantified by the molar ratio of ATP/(ATP + ADP). A 35% decrease in cellular energy is shown to occur in vivo upon a nutrient downshift that gives rise to cyst formation. When this energy decline is replicated in vitro, the phosphorylation level of CheS3 is reduced by ~75%. Finally, we also show that ADP-mediated reduction of CheS3 phosphorylation is a consequence of ADP enhancing autodephosphorylation of CheS3.

Rhodospirillum centenum is a Gram-negative alphaproteobacterium that is capable of differentiating into dormant cysts that are metabolically inactive and desiccation resistant. Like spores synthesized by many Gram-positive species, dormant R. centenum cysts germinate in response to an environmental signal, indicating that conditions favor survival and proliferation. Factors that induce germination are called germinants and are often both niche and species specific. In this study, we have identified photosynthesis as a niche-specific germinant for R. centenum cyst germination. Specifically, excitation of wild-type cysts suspended in a nutrient-free buffer with far-red light at >750 nm results in rapid germination. This is in stark contrast to mutant strains deficient in photosynthesis that fail to germinate upon exposure to far-red light under all assayed conditions. We also show that photosynthesis-induced germination occurs in a carbon- and nitrogen-free buffer even in strains that are deficient in carbon or nitrogen fixation. These results demonstrate that photosynthesis not only is necessary for germination but is itself sufficient for the germination of R. centenum cysts. IMPORTANCE Environmental cues that signal Gram-positive spores to germinate (termed germinants) have been identified for several Bacillus and Clostridium species. These studies showed that germinants are niche and species specific. For example, Clostridium difficile spores sense bile salts as a germinant as their presence informs these cells of an intestinal environment. Bacillus fastidiosus spores use uric acid as a germinant that is present in soil and poultry litter as this species inhabits poultry litter. It is evident from these studies that dormant cells sample their environment to assess whether conditions are advantageous for the propagation and survival of vegetative cells. To date, a limited number of germinants have been defined for only a few Gram-positive spore-forming species. Beyond that group, there is scant information on what cues signal dormant cells to exit dormancy. In our study, we show that the versatile Gram-negative photosynthetic bacterium Rhodospirillum centenum uses light-driven photosynthesis, and not the availability of nutrients, to trigger the germination of dormant cysts. This use of light-driven photosynthesis as a germinant is surprising as this species is also capable of growing under dark conditions using exogenous carbon sources for energy. Consequently, photosynthetic growth appears to be the preferred growth mechanism by this species.