Phototrophic microorganisms are promising resources for green biotechnology. Compared to heterotrophic microorganisms, however, the cellular economy of phototrophic growth is still insufficiently understood. We provide a quantitative analysis of light-limited, light-saturated, and light-inhibited growth of the cyanobacterium Synechocystis sp. PCC 6803 using a reproducible cultivation setup. We report key physiological parameters, including growth rate, cell size, and photosynthetic activity over a wide range of light intensities. Intracellular proteins were quantified to monitor proteome allocation as a function of growth rate. Among other physiological acclimations, we identify an upregulation of the translational machinery and downregulation of light harvesting components with increasing light intensity and growth rate. The resulting growth laws are discussed in the context of a coarse-grained model of phototrophic growth and available data obtained by a comprehensive literature search. Our insights into quantitative aspects of cyanobacterial acclimations to different growth rates have implications to understand and optimize photosynthetic productivity.

In living cells, the perception of environmental stress and the subsequent transduction of stress signals are primary events in the acclimation to changes in the environment. Some molecular sensors and transducers of environmental stress cannot be identified by traditional and conventional methods. Based on genomic information, a systematic approach has been applied to the solution of this problem in cyanobacteria, involving mutagenesis of potential sensors and signal transducers in combination with DNA microarray analyses for the genome-wide expression of genes. Forty-five genes for the histidine kinases (Hiks), 12 genes for serine-threonine protein kinases (Spks), 42 genes for response regulators (Rres), seven genes for RNA polymerase sigma factors, and nearly 70 genes for transcription factors have been successfully inactivated by targeted mutagenesis in the unicellular cyanobacterium Synechocystis sp. PCC 6803. Screening of mutant libraries by genome-wide DNA microarray analysis under various stress and non-stress conditions has allowed identification of proteins that perceive and transduce signals of environmental stress. Here we summarize recent progress in the identification of sensory and regulatory systems, including Hiks, Rres, Spks, sigma factors, transcription factors, and the role of genomic DNA supercoiling in the regulation of the responses of cyanobacterial cells to various types of stress.

Changes in the supercoiling of genomic DNA play an important role in the regulation of gene expression. We compared the genome-wide expression of genes in cells of the cyanobacterium Synechocystis sp. PCC 6803 when they were subjected to salt, cold, and heat stress, in the presence of novobiocin, an inhibitor of DNA gyrase, and in its absence. The analysis revealed that the expression of a large number of stress-inducible genes depends on the extent of genomic DNA supercoiling. The function of the two-component regulatory systems, which are known as sensors and transducers of salt, cold, and heat stress, depends on, and might be controlled by, the degree of supercoiling of the genomic DNA. These results suggest that stress-induced changes in superhelicity of genomic DNA provide an important permissive background for successful acclimatization of cyanobacterial cells to stress conditions.

Serine/threonine protein kinases (STPKs) are the major participants in intracellular signal transduction in eukaryotes, such as yeasts, fungi, plants, and animals. Genome sequences indicate that these kinases are also present in prokaryotes, such as cyanobacteria. However, their roles in signal transduction in prokaryotes remain poorly understood. We have attempted to identify the roles of STPKs in response to heat stress in the prokaryotic cyanobacterium Synechocystis sp. PCC 6803, which has 12 genes for STPKs. Each gene was individually inactivated to generate a gene-knockout library of STPKs. We applied in vitro Ser/Thr protein phosphorylation and phosphoproteomics and identified the methionyl-tRNA synthetase, large subunit of RuBisCO, 6-phosphogluconate dehydrogenase, translation elongation factor Tu, heat-shock protein GrpE, and small chaperonin GroES as the putative targets for Ser/Thr phosphorylation. The expressed and purified GroES was used as an external substrate to screen the protein extracts of the individual mutants for their Ser/Thr kinase activities. The mutants that lack one of the three protein kinases, SpkC, SpkF, and SpkK, were unable to phosphorylate GroES in vitro, suggesting possible interactions between them towards their substrate. Complementation of the mutated SpkC, SpkF, and SpkK leads to the restoration of the ability of cells to phosphorylate the GroES. This suggests that these three STPKs are organized in a sequential order or a cascade and they work one after another to finally phosphorylate the GroES.

Phototrophic microorganisms are promising resources for green biotechnology. 13 Compared to heterotrophic microorganisms, however, the cellular economy of phototrophic 14 growth is still insufficiently understood. We provide a quantitative analysis of light-limited, 15 light-saturated, and light-inhibited growth of the cyanobacterium Synechocystis sp. PCC 6803 using 16 a reproducible cultivation setup. We report key physiological parameters, including growth rate, cell 17 size, and photosynthetic activity over a wide range of light intensities. Intracellular proteins were 18 quantified to monitor proteome allocation as a function of growth rate. Among other physiological 19 adaptations, we identify an upregulation of the translational machinery and downregulation of light 20 harvesting components with increasing light intensity and growth rate. The resulting growth laws 21 are discussed in the context of a coarse-grained model of phototrophic growth and available data 22 obtained by a comprehensive literature search. Our insights into quantitative aspects of 23 cyanobacterial adaptations to different growth rates have implications to understand and optimize 24 photosynthetic productivity. 25 26 2014). While quantitative insight into the cellular economy of phototrophic microorganisms is 36 still scarce, the cellular economy of heterotrophic growth has been studied extensively-starting 37 with the seminal works of Monod, Neidhardt, and others (Neidhardt et al., 1990; Neidhardt, 1999; 38 Jun et al., 2018) to more recent quantitative studies of microbial resource allocation (Molenaar 39 et al.40 Maitra and Dill, 2015; Weiße et al., 2015). In response to changing environments, heterotrophic 41 1 of 28 Manuscript submitted to eLife microorganisms are known to differentially allocate their resources: with increasing growth rate, 42 heterotrophic microorganisms typically exhibit upregulation of ribosomes and other proteins 43 related to translation and protein synthesis (Scott et al., 2010; Molenaar et al., 2009; Peebo et al., 44 2015), exhibit complex changes in transcription profiles, e.g. (Klumpp et al., 2009; Matsumoto et al., 45 2013), and increase cell size (Kafri et al., 2016). The molecular limits of heterotrophic growth have 46 been described thoroughly (Kafri et al., 2016; Erickson et al., 2017; Scott et al., 2014; Metzl-Raz 47 et al., 2017; Klumpp et al., 2013). 48 In contrast, only few studies so far have addressed the limits of cyanobacterial growth from an 49 experimental perspective (Bernstein et al., 2016; Yu et al., 2015; Abernathy et al., 2017; Ungerer 50 et al., 2018; Jahn et al., 2018). Of particular interest were the adaptations that enable fast pho-51 toautotrophic growth (Bernstein et al., 2016; Yu et al., 2015; Abernathy et al., 2017; Ungerer et al., 52 2018). The cyanobacterium with the highest known photoautotrophic growth rate, growing with a 53 doubling time of up to ∼ 1.5h, is the strain Synechococcus elongatus UTEX 2973 (Ungerer et al., 54 2018). Compared to its c...