Costs of Clock-Environment Misalignment in Individual Cyanobacterial Cells

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130

Cyanobacteria are an integral part of Earth's biogeochemical cycles and a promising resource for the synthesis of renewable bioproducts from atmospheric CO 2 . Growth and metabolism of cyanobacteria are inherently tied to the diurnal rhythm of light availability. As yet, however, insight into the stoichiometric and energetic constraints of cyanobacterial diurnal growth is limited. Here, we develop a computational framework to investigate the optimal allocation of cellular resources during diurnal phototrophic growth using a genome-scale metabolic reconstruction of the cyanobacterium Synechococcus elongatus PCC 7942. We formulate phototrophic growth as an autocatalytic process and solve the resulting time-dependent resource allocation problem using constraint-based analysis. Based on a narrow and well-defined set of parameters, our approach results in an ab initio prediction of growth properties over a full diurnal cycle. The computational model allows us to study the optimality of metabolite partitioning during diurnal growth. The cyclic pattern of glycogen accumulation, an emergent property of the model, has timing characteristics that are in qualitative agreement with experimental findings. The approach presented here provides insight into the time-dependent resource allocation problem of phototrophic diurnal growth and may serve as a general framework to assess the optimality of metabolic strategies that evolved in phototrophic organisms under diurnal conditions. constraint-based analysis | whole-cell models | bioenergetics | metabolism | circadian clock C yanobacterial photoautotrophic growth requires a highly coordinated distribution of cellular resources to different intracellular processes, including the de novo synthesis of proteins, ribosomes, lipids, and other cellular components. For unicellular organisms, the optimal allocation of limiting resources is a key determinant of evolutionary fitness. Owing to the importance of cellular resource allocation for understanding evolutionary trade-offs in bacterial metabolism, the cellular "protein economy" and its implications for bacterial growth laws have been studied extensively, albeit almost exclusively for heterotrophic organisms under stationary environmental conditions (1-7). For photoautotrophic organisms, including cyanobacteria, growthdependent resource allocation is further subject to diurnal lightdark (LD) cycles that partition cellular metabolism into distinct phases. Recent experimental results have demonstrated the relevance of time-specific synthesis for cellular survival and growth (8-10). Nonetheless, the implications and consequences of a diurnal environment for the cellular resource allocation problem are insufficiently understood, and computational approaches hitherto developed for heterotrophic growth are not straightforwardly applicable to diurnal phototrophic growth (11).Here, we propose a computational framework to quantitatively assess the optimality of diurnal resource allocation for phototrophic growth. We are primarily interested i...

Section: Discussionmentioning

confidence: 79%

mentioning

confidence: 99%

Cellular trade-offs and optimal resource allocation during cyanobacterial diurnal growth

Reimers

Knoop

Bockmayr

et al. 2017

104

130

“…Places where the CI does not overlap indicate a statistically significant difference in the model. (52), as well as data indicating that the fitness advantage conferred by the S. elongatus circadian clock is more likely due to the ability to anticipate a coming dark period rather than a morning period (53).…”

Section: Discussionmentioning

confidence: 99%

Redox crisis underlies conditional light–dark lethality in cyanobacterial mutants that lack the circadian regulator, RpaA

Diamond

Rubin

Shultzaberger

et al. 2017

Cyanobacteria evolved a robust circadian clock, which has a profound influence on fitness and metabolism under daily lightdark (LD) cycles. In the model cyanobacterium Synechococcus elongatus PCC 7942, a functional clock is not required for diurnal growth, but mutants defective for the response regulator that mediates transcriptional rhythms in the wild-type, regulator of phycobilisome association A (RpaA), cannot be cultured under LD conditions. We found that rpaA-null mutants are inviable after several hours in the dark and compared the metabolomes of wild-type and rpaA-null strains to identify the source of lethality. Here, we show that the wild-type metabolome is very stable throughout the night, and this stability is lost in the absence of RpaA. Additionally, an rpaA mutant accumulates excessive reactive oxygen species (ROS) during the day and is unable to clear it during the night. The rpaA-null metabolome indicates that these cells are reductant-starved in the dark, likely because enzymes of the primary nighttime NADPH-producing pathway are direct targets of RpaA. Because NADPH is required for processes that detoxify ROS, conditional LD lethality likely results from inability of the mutant to activate reductant-requiring pathways that detoxify ROS when photosynthesis is not active. We identified second-site mutations and growth conditions that suppress LD lethality in the mutant background that support these conclusions. These results provide a mechanistic explanation as to why rpaA-null mutants die in the dark, further connect the clock to metabolism under diurnal growth, and indicate that RpaA likely has important unidentified functions during the day.C yanobacteria are both key agents of global carbon and nitrogen cycles and promising platforms for renewable chemicals, fuels, and nutraceuticals (1-3). Understanding the control mechanisms that govern the flow of carbon and nitrogen through these organisms is crucial for predicting their behavior in natural environments as well as for improving engineering strategies. Although the basic pathways for carbon and nitrogen metabolism, and their regulation, are well understood in heterotrophic bacteria, cyanobacteria exhibit important deviations in these core metabolic pathways (4-7). Additionally, metabolic control mechanisms in cyanobacteria evolved to be compatible with photoautotrophic metabolism and the dramatic shifts that are imposed on those pathways by predictable daily light-dark (LD) cycles. Examples include enzymatic activity that responds to light-dependent cellular redox changes (8-11); the preference for NADPH, the reductant produced by the photochemical reactions, over NADH by many biosynthetic enzymes (12, 13); and a circadian clock that drives 24-h transcriptional rhythms in most genes (14-16).A daily LD cycle presents a strong metabolic driver for the photosynthetic cyanobacteria, but a circadian clock also imposes daily cycles in transcription and redox regulatory systems (17, 18). Circadian measurements historically have been performed in...

“…Note that the resonance period was ∼30 h in this study. The resonance period may depend on experimental conditions such as the concentrations of Kai proteins, and clocks in vivo may have a resonance period closer to 24 h. Further, the concept of circadian resonance was proposed in the context of enhanced growth and metabolic activity, when the period of light/dark cycles is close to that of the endogenous circadian clock (42)(43)(44). The acquisition of an optimal phase relationship between the clock and environmental rhythms when the periods are close may provide an explanation.…”

Section: Discussionmentioning

confidence: 99%

Low temperature nullifies the circadian clock in cyanobacteria through Hopf bifurcation

Murayama

Kori

Oshima

et al. 2017

Cold temperatures lead to nullification of circadian rhythms in many organisms. Two typical scenarios explain the disappearance of rhythmicity: the first is oscillation death, which is the transition from self-sustained oscillation to damped oscillation that occurs at a critical temperature. The second scenario is oscillation arrest, in which oscillation terminates at a certain phase. In the field of nonlinear dynamics, these mechanisms are called the Hopf bifurcation and the saddle-node on an invariant circle bifurcation, respectively. Although these mechanisms lead to distinct dynamical properties near the critical temperature, it is unclear to which scenario the circadian clock belongs. Here we reduced the temperature to dampen the reconstituted circadian rhythm of phosphorylation of the recombinant cyanobacterial clock protein KaiC. The data led us to conclude that Hopf bifurcation occurred at ∼19°C. Below this critical temperature, the self-sustained rhythms of KaiC phosphorylation transformed to damped oscillations, which are predicted by the Hopf bifurcation theory. Moreover, we detected resonant oscillations below the critical temperature when temperature was periodically varied, which was reproduced by numerical simulations. Our findings suggest that the transition to a damped oscillation through Hopf bifurcation contributes to maintaining the circadian rhythm of cyanobacteria through resonance at cold temperatures.