Changes in Transcript Abundance in Chlamydomonas reinhardtii following Nitrogen Deprivation Predict Diversion of Metabolism

Abstract

Like many microalgae, Chlamydomonas reinhardtii forms lipid droplets rich in triacylglycerols when nutrient deprived. To begin studying the mechanisms underlying this process, nitrogen (N) deprivation was used to induce triacylglycerol accumulation and changes in developmental programs such as gametogenesis. Comparative global analysis of transcripts under induced and noninduced conditions was applied as a first approach to studying molecular changes that promote or accompany triacylglycerol accumulation in cells encountering a new nutrient environment. Towards this goal, high-throughput sequencing technology was employed to generate large numbers of expressed sequence tags of eight biologically independent libraries, four for each condition, N replete and N deprived, allowing a statistically sound comparison of expression levels under the two tested conditions. As expected, N deprivation activated a subset of control genes involved in gametogenesis while down-regulating protein biosynthesis. Genes for components of photosynthesis were also down-regulated, with the exception of the PSBS gene. N deprivation led to a marked redirection of metabolism: the primary carbon source, acetate, was no longer converted to cell building blocks by the glyoxylate cycle and gluconeogenesis but funneled directly into fatty acid biosynthesis. Additional fatty acids may be produced by membrane remodeling, a process that is suggested by the changes observed in transcript abundance of putative lipase genes. Inferences on metabolism based on transcriptional analysis are indirect, but biochemical experiments supported some of these deductions. The data provided here represent a rich source for the exploration of the mechanism of oil accumulation in microalgae.

Chlamydomonas reinhardtii PsbS Protein Is Functional and Accumulates Rapidly and Transiently under High Light

“…Even if PsbS proteins in Chlamydomonas have not been detected in previous analyses, the presence of conserved psbS genes, their expression under nitrogen deprivation conditions (Miller et al, 2010), and the expression of the genes during a dark to light shift as found in a very recent report (Zones et al, 2015) suggest a role for PsbS in the alga. Thus, in this work, we have reinvestigated the expression and accumulation of PsbS in Chlamydomonas during light stress and other stress conditions in order to determine whether this protein could play a role in NPQ under particular conditions and compared our results with those obtained on LhcSR3.…”

“…It has been reported that under nitrogen deprivation, psbs mRNA accumulates to high levels (Miller et al, 2010). Thus, we performed immunoblot experiments to check the accumulation of the PsbS protein in cells grown in these conditions.…”

The Response of Diatom Central Carbon Metabolism to Nitrogen Starvation Is Different from That of Green Algae and Higher Plants

“…In higher plants, this is usually in the form of starch (Diaz et al, 2005;Wingler et al, 2006), and Peng et al (2007) described increased transcript levels of two genes involved in starch biosynthesis in Arabidopsis (Arabidopsis thaliana) under nitrogen starvation. In the fresh water green alga Chlamydomonas reinhardtii, on the other hand, excess carbon is reportedly directed to fatty acid biosynthesis (Wang et al, 2009;Moellering and Benning, 2010), and Miller et al (2010) found increased expression of genes involved in lipid biosynthesis in C. reinhardtii when nitrogen was not available. Diatoms are known to store carbon as chrysolaminaran (a b-1,3-glucan) or as lipids (Armbrust et al, 2004;Kroth et al, 2008); however, proteins directly related to the synthesis or degradation of these compounds were not identified in this study.…”

“…In addition, in both Arabidopsis (Peng et al, 2007) and C. reinhardtii (Miller et al, 2010), transcript levels of genes associated with the TCA cycle either decreased in abundance or remained unchanged in response to nitrogen deprivation ( Fig. 7; Supplemental Table S2).…”

“…A, T. pseudonana, grown under conditions comparable to this study . B, C. reinhardtii, 48 h after the transfer of cells from 10 mM ammonium to nitrogen-free medium (Miller et al, 2010). C, Arabidopsis, at the onset of nitrogen limitation in plants grown with 3 mM nitrate (Peng et al, 2007).…”