The response of Chlamydomonas reinhardtii to nitrogen deprivation: a systems biology analysis

Abstract

SUMMARYDrastic alteration in macronutrients causes large changes in gene expression in the photosynthetic unicellular alga Chlamydomonas reinhardtii. Preliminary data suggested that cells follow a biphasic response to this change hinging on the initiation of lipid accumulation, and we hypothesized that drastic repatterning of metabolism also followed this biphasic modality. To test this hypothesis, transcriptomic, proteomic, and metabolite changes that occur under nitrogen (N) deprivation were analyzed. Eight sampling times were selected covering the progressive slowing of growth and induction of oil synthesis between 4 and 6 h after N deprivation. Results of the combined, systems-level investigation indicated that C. reinhardtii cells sense and respond on a large scale within 30 min to a switch to N-deprived conditions turning on a largely gluconeogenic metabolic state, which then transitions to a glycolytic stage between 4 and 6 h after N depletion. This nitrogen-sensing system is transduced to carbon-and nitrogen-responsive pathways, leading to down-regulation of carbon assimilation and chlorophyll biosynthesis, and an increase in nitrogen metabolism and lipid biosynthesis. For example, the expression of nearly all the enzymes for assimilating nitrogen from ammonium, nitrate, nitrite, urea, formamide/acetamide, purines, pyrimidines, polyamines, amino acids and proteins increased significantly. Although arginine biosynthesis enzymes were also rapidly up-regulated, arginine pool size changes and isotopic labeling results indicated no increased flux through this pathway.

The Regulation of Photosynthetic Structure and Function during Nitrogen Deprivation in Chlamydomonas reinhardtii

“…The false-positive rate of protein identification was assessed with an automatic decoy database search, and identified proteins were grouped based on 98% homology with Scaffold version 3.1 (Proteome Software). Full details can be found in Park et al (2014).…”

“…reinhardtii samples were harvested at 0, 0.5, 1, 2, 4, 6, 12, and 24 h after N deprivation, and transcripts were analyzed by high-throughput sequencing as described by Park et al (2014). Nitrogen-replete cells (time 0) were used as the reference for all subsequent time points.…”

“…Transcript analysis was conducted as described in Park et al (2014), which includes the mapping, parameters for single nucleotide polymorphism analysis, statistical data tools, and functional annotations. Briefly, cells were harvested by centrifugation at 0°C, resuspended in RNAlater solution (Qiagen Sciences), and stored at between -80°C and -60°C ; RNA was extracted from two biological replicates for each time point using TRIzol reagent (Invitrogen) according to the manufacturer's protocol.…”

Algal Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase, Triacylglycerol Accumulation Regulator1, Regulates Accumulation of Triacylglycerol in Nitrogen or Sulfur Deficiency

“…wild-type, and C1 lines were estimated by the DESeq algorithm with a q-value cutoff of less than 0.05 from RNA sequencing (RNA-seq) data (Supplemental Table S1; Supplemental Data Set S1). For RNA-seq analysis, RNA was extract from each line 6 h after being transferred to Tris-acetate phosphate medium without a nitrogen source (TAP 2 N; normal TAP medium as a control), because the previous reports revealed that TAG accumulation starts in the cells (Park et al, 2015); expression of a large part of conserved N-regulated genes has already responded (Schmollinger et al, 2014) at that time point. Consequently, the expression levels of 937, 509, and 405 genes were increased significantly in N-deficient conditions in tar1-1, wild-type, and C1 cells, respectively (Fig.…”

Experimentally Induced Bleaching in the Sea Anemone Exaiptasia Supports Glucose as a Main Metabolite Associated with Its Symbiosis

“…G6PDH is the first step in the pentose phosphate pathway that regenerates NADPH for anabolic metabolism, as well as pentoses like ribose-5 phosphate, a precursor in the synthesis of nucleotides. When increased, the level of enzymes involved in oxidative pentose phosphate pathway (e.g., G6PDH and 6-P gluconate dehydrogenase), rather than leading to the accumulation of carbon skeletons for cellular growth, regenerates NADPH, essential for the reduction in fatty acid biosynthesis [34].…”