Triacylglycerol (TAG) is a storage lipid used for food purposes and as a renewable feedstock for biodiesel production. WRINKLED1 (WRI1) is a transcription factor that governs fatty acid (FA) synthesis and, indirectly, TAG accumulation in oil-storing plant tissues, and its ectopic expression has led to TAG accumulation in vegetative tissues of different dicotyledonous plants. The ectopic expression of BdWRI1 in the grass Brachypodium distachyon induced the transcription of predicted genes involved in glycolysis and FA biosynthesis, and TAG content was increased up to 32.5-fold in 8-week-old leaf blades. However, the ectopic expression of BdWRI1 also caused cell death in leaves, which has not been observed previously in dicotyledonous plants such as Arabidopsis (Arabidopsis thaliana). Lipid analysis indicated that the free FA content was 2-fold elevated in BdWRI1-expressing leaf blades of B. distachyon. The transcription of predicted genes involved in b-oxidation was induced. In addition, linoleic FA treatment caused cell death in B. distachyon leaf blades, an effect that was reversed by the addition of the FA biosynthesis inhibitor cerulenin. Taken together, ectopic expression of BdWRI1 in B. distachyon enhances FA biosynthesis and TAG accumulation in leaves, as expected, but also leads to increased free FA content, which has cytotoxic effects leading to cell death. Thus, while WRI appears to ubiquitously affect FA biosynthesis and TAG accumulation in diverse plants, its ectopic expression can lead to undesired side effects depending on the context of the specific lipid metabolism of the respective plant species.

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Under nitrogen (N) starvation, microalgae increase carbon storage in the form of lipid droplets while also downregulating photosynthesis and eventually terminating growth. To improve lipid yield, we asked whether lipid droplets and N starvation responses can be induced without limiting growth or photosynthesis. In the chlorophyte Chlamydomonas reinhardtii, gametogenesis is induced either by N starvation or by growth with arginine as the sole N source. We showed that arginine cultures supported robust phototrophic growth, constitutively turned on N starvation-induced genes, and increased lipid droplets. The lipids accumulated in arginine cultures exhibited strong enrichment of saturated and monounsaturated fatty acids, a preferred characteristic of biodiesel precursors. The diatom Phaeodactylum tricornutum also accumulated lipid droplets in arginine culture without growth impairment. We document a system wherein N starvation responses are induced without compromising photosynthesis or growth, thereby suited to the producing valuable chemicals and biofuel precursors without requiring stressors in microalgae.