Ammonia-oxidizing archaea (AOA) are among the most abundant microorganisms and key players in the global nitrogen and carbon cycles. They share a common energy metabolism but represent a heterogeneous group with respect to their environmental distribution and adaptions, growth requirements, and genome contents. We report here the genome and proteome of Nitrososphaera viennensis EN76, the type species of the archaeal class Nitrososphaeria of the phylum Thaumarchaeota encompassing all known AOA. N. viennensis is a soil organism with a 2.52-Mb genome and 3,123 predicted proteincoding genes. Proteomic analysis revealed that nearly 50% of the predicted genes were translated under standard laboratory growth conditions. Comparison with genomes of closely related species of the predominantly terrestrial Nitrososphaerales as well as the more streamlined marine Nitrosopumilales [Candidatus (Ca.) order] and the acidophile "Ca. Nitrosotalea devanaterra" revealed a core genome of AOA comprising 860 genes, which allowed for the reconstruction of central metabolic pathways common to all known AOA and expressed in the N. viennensis and "Ca. Nitrosopelagicus brevis" proteomes. Concomitantly, we were able to identify candidate proteins for as yet unidentified crucial steps in central metabolisms. In addition to unraveling aspects of core AOA metabolism, we identified specific metabolic innovations associated with the Nitrososphaerales mediating growth and survival in the soil milieu, including the capacity for biofilm formation, cell surface modifications and cell adhesion, and carbohydrate conversions as well as detoxification of aromatic compounds and drugs. ammonia oxidation | proteomics | archaea | comparative genomics | biofilm

BackgroundNitrogen starvation is known to cause drastic alterations in physiology and metabolism leading to the accumulation of lipid bodies in many microalgae, and it thus presents an important alternative for biofuel production. However, despite the importance of this process, the molecular mechanisms that mediate the metabolic remodeling induced by N starvation and especially by stress recovery are still poorly understood, and new candidates for bioengineering are needed to make this process useful for biofuel production.ResultsWe have studied the molecular changes involved in the adaptive mechanisms to N starvation and full recovery of the vegetative cells in the microalga Chlamydomonas reinhardtii during a four-day time course.High throughput mass spectrometry was employed to integrate the proteome and the metabolome with physiological changes. N starvation led to an accumulation of oil bodies and reduced Fv/Fm.. Distinct enzymes potentially participating in the carbon-concentrating mechanism (CAH7, CAH8, PEPC1) are strongly accumulated. The membrane composition is changed, as indicated by quantitative lipid profiles. A reprogramming of protein biosynthesis was observed by increased levels of cytosolic ribosomes, while chloroplastidic were dramatically reduced. Readdition of N led to, the identification of early responsive proteins mediating stress recovery, indicating their key role in regaining and sustaining normal vegetative growth.Analysis of the data with multivariate correlation analysis, Granger causality, and sparse partial least square (sPLS) provided a functional network perspective of the molecular processes. Cell growth and N metabolism were clearly linked by the branched chain amino acids, suggesting an important role in this stress. Lipid accumulation was also tightly correlated to the COP II protein, involved in vesicle and lysosome coating, and a major lipid droplet protein. This protein, together with other key proteins mediating signal transduction and adaption (BRI1, snRKs), constitute a series of new metabolic and regulatory targets.ConclusionsThis work not only provides new insights and corrects previous models by analyzing a complex dataset, but also increases our biochemical understanding of the adaptive mechanisms to N starvation in Chlamydomonas, pointing to new bioengineering targets for increased lipid accumulation, a key step for a sustainable and profitable microalgae-based biofuel production.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-014-0171-1) contains supplementary material, which is available to authorized users.

Chlamydomonas reinhardtii is one of the most important model organisms nowadays phylogenetically situated between higher plants and animals (Merchant et al. 2007). Stress adaptation of this unicellular model algae is in the focus because of its relevance to biomass and biofuel production. Here, we have studied cold stress adaptation of C. reinhardtii hitherto not described for this algae whereas intensively studied in higher plants. Toward this goal, high throughput mass spectrometry was employed to integrate proteome, metabolome, physiological and cell-morphological changes during a time-course from 0 to 120 h. These data were complemented with RT-qPCR for target genes involved in central metabolism, signaling, and lipid biosynthesis. Using this approach dynamics in central metabolism were linked to cold-stress dependent sugar and autophagy pathways as well as novel genes in C. reinhardtii such as CKIN1, CKIN2 and a hitherto functionally not annotated protein named CKIN3. Cold stress affected extensively the physiology and the organization of the cell. Gluconeogenesis and starch biosynthesis pathways are activated leading to a pronounced starch and sugar accumulation. Quantitative lipid profiles indicate a sharp decrease in the lipophilic fraction and an increase in polyunsaturated fatty acids suggesting this as a mechanism of maintaining membrane fluidity. The proteome is completely remodeled during cold stress: specific candidates of the ribosome and the spliceosome indicate altered biosynthesis and degradation of proteins important for adaptation to low temperatures. Specific proteasome degradation may be mediated by the observed cold-specific changes in the ubiquitinylation system. Sparse partial least squares regression analysis was applied for protein correlation network analysis using proteins as predictors and Fv/Fm, FW, total lipids, and starch as responses. We applied also Granger causality analysis and revealed correlations between proteins and metabolites otherwise not detectable. Twenty percent of the proteins responsive to cold are uncharacterized proteins. This presents a considerable resource for new discoveries in cold stress biology in alga and plants. Molecular &