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Whole cell response to nitrogen deprivation in the diatom Phaeodactylum tricornutum

ABSTRACT: We combined transcriptional and metabolite analyses to monitor the effect of N deficiency at different molecular levels in order to get better insight on the acclimation strategies P. tricornutum employs under nitrogen limitation. Physiological data like neutral lipid measurement, cell growth and other cell chemistry measurements further complemented our molecular data. We showed that P. tricornutum is able to remobilize nitrogen through catabolism of various internal nitrogen-containing resources such as amino acids and proteins. N starvation was also accompanied by reduction in pigment pools as well as photosynthetic capacity. The global expression data showed that majority of metabolic changes were related to carbon and lipid metabolism. Decreased levels of carbon skeletons due to suppression of the Calvin cycle was compensated by breakdown of chrysolaminaran leading to up-regulation of OPPP, glycolysis, pyruvate metabolism and TCA cycle. These pathways provide precursors for fatty acid biosynthesis. In addition, remodeling of membrane lipids and up-regulation of the de novo TAG biosynthetic pathway was further supported by physiological measurements of neutral lipids, indicating TAG accumulation under N limitation. Our study gives a detailed image of adaptation of P. tricornutum to N starvation, and can be used for future metabolic manipulations to increase TAG production.

ORGANISM(S): Phaeodactylum tricornutum

PROVIDER: GSE58946 | GEO | 2015/06/01

SECONDARY ACCESSION(S): PRJNA253958

REPOSITORIES: GEO

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Project description:The oleaginous green microalga Neochloris oleoabundans accumulates both starch and lipids to high levels under stress conditions such as nitrogen starvation. In order to steer the metabolism towards starch or lipids only, it is important to understand the mechanisms behind it. In this study physiological changes and gene expression upon nitrogen starvation were analysed in controlled flat-panel photobioreactors over both short- and long-term time-scales. Starch accumulation is transient and occurs rapidly within 24 hrs upon starvation, while lipid accumulation lasts longer and reaches a maximum after 4 days. The major fraction of accumulated lipids is composed of de novo synthesized neutral lipids - triacylglycerides (TAG) - and is characterized by a decreased composition of the polyunsaturated fatty acids (PUFAs) C18:3 and C16:3 and an increased composition of the mono-unsaturated and saturated fatty acids C18:1/C16:1 and C18:0/C16:0, respectively. RNA-sequencing revealed that genes related to starch biosynthesis and degradation show different temporal expression dynamics compared to those of lipid biosynthesis. An immediate rapid increase in starch synthesis gene expression is followed by an increase in starch degradation and decrease in starch synthesis gene expression. In contrast, increased gene expression for fatty acid and TAG synthesis is initiated later and occurs more gradually. Expression of several fatty acid desaturase (FAD) genes was decreased upon starvation, which corresponds to the observed changes to higher levels of mono-unsaturated and saturated fatty acids. Moreover, several homologs of transcription regulators that were implicated in controlling starch and lipid metabolism in other microalgae showed differential gene expression and might be key regulators of starch and lipid metabolism in N. oleoabundans as well. Promising candidates for future metabolic engineering are a DYRKP homolog that in Chlamydomonas acts as a negative regulator of carbon storage and photosynthetic efficiency under N-starvation, and two bZIP-type regulators that have been implicated to control several steps in microalgal TAG synthesis. Our data for the first time show the temporal dynamics of storage compound accumulation and transcriptional changes on a short- and long-term time-scale during nitrogen starvation in N. oleoabundans, and increases insight into the genetic foundation for starch and lipid metabolism in this microalga. This information and the identified target genes can now be used for metabolic engineering strategies towards tailored N. oleoabundans strains for industrial applications with increased lipid production and altered fatty acid composition.

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Whole cell response to nitrogen deprivation in the diatom Phaeodactylum tricornutum

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