Summary: The accumulation of storage compounds during seed development ensures the survival of the young seedling, and also provides nutrition to humans and animals in the form of foods and feeds. The putative AP2/EREBP transcription factor WRINKLED1 (WRI1) is involved in the regulation of seed storage metabolism in Arabidopsis. A splicing mutant allele, wri1‐1, caused the reduction of seed oil accumulation. Glycolysis was compromised in this mutant, rendering developing embryos unable to efficiently convert sucrose into precursors of triacylglycerol biosynthesis. Expression of the WRINKLED1 cDNA under the control of the cauliflower mosaic virus 35S‐promoter led to increased seed oil content. Moreover, the ectopic expression of the WRINKLED1 cDNA caused the accumulation of triacylglycerols in developing seedlings. This effect depended upon the presence of glucose in the growth medium or other sugars readily metabolized to glucose. Oil‐accumulating seedlings showed aberrant development consistent with a prolonged embryonic state.

Summary: Triacylglycerols produced by plants are one of the most energy‐rich and abundant forms of reduced carbon available from nature. Given their chemical similarities, plant oils represent a logical substitute for conventional diesel, a non‐renewable energy source. However, as plant oils are too viscous for use in modern diesel engines, they are converted to fatty acid esters. The resulting fuel is commonly referred to as biodiesel, and offers many advantages over conventional diesel. Chief among these is that biodiesel is derived from renewable sources. In addition, the production and subsequent consumption of biodiesel results in less greenhouse gas emission compared to conventional diesel. However, the widespread adoption of biodiesel faces a number of challenges. The biggest of these is a limited supply of biodiesel feedstocks. Thus, plant oil production needs to be greatly increased for biodiesel to replace a major proportion of the current and future fuel needs of the world. An increased understanding of how plants synthesize fatty acids and triacylglycerols will ultimately allow the development of novel energy crops. For example, knowledge of the regulation of oil synthesis has suggested ways to produce triacylglycerols in abundant non‐seed tissues. Additionally, biodiesel has poor cold‐temperature performance and low oxidative stability. Improving the fuel characteristics of biodiesel can be achieved by altering the fatty acid composition. In this regard, the generation of transgenic soybean lines with high oleic acid content represents one way in which plant biotechnology has already contributed to the improvement of biodiesel.

An allelic series of the novel argonaute mutant ( ago1‐1 to ago1‐6 ) of the herbaceous plant Arabidopsis thaliana has been isolated. The ago1 mutation pleotropically affects general plant architecture. The apical shoot meristem generates rosette leaves and a single stem, but axillary meristems rarely develop. Rosette leaves lack a leaf blade but still show adaxial/abaxial differentiation. Instead of cauline leaves, filamentous structures without adaxial/abaxial differentiation develop along the stem and an abnormal inflorescence bearing infertile flowers with filamentous organs is produced. Two independent T‐DNA insertions into the AGO1 locus led to the isolation of two corresponding genomic sequences as well as a complete cDNA. The AGO1 locus was mapped close to the marker mi291a on chromosome 1. Antisense expression of the cDNA resulted in a partial mutant phenotype. Sense expression caused some transgenic lines to develop goblet‐like leaves and petals. The cDNA encodes a putative 115 kDa protein with sequence similarity to translation products of a novel gene family present in nematodes as well as humans. No specific function has been assigned to these genes. Similar proteins are not encoded by the genomes of yeast or bacteria, suggesting that AGO1 belongs to a novel class of genes with a function specific to multicellular organisms.

During oil deposition in developing seeds of Arabidopsis, photosynthate is imported in the form of carbohydrates into the embryo and converted to triacylglycerols. To identify genes essential for this process and to investigate the molecular basis for the developmental regulation of oil accumulation, mutants producing wrinkled, incompletely filled seeds were isolated. A novel mutant locus, wrinkled1 (wri1), which maps to the bottom of chromosome 3 and causes an 80% reduction in seed oil content, was identified. Wild-type and homozygouswri1 mutant plantlets or mature plants were indistinguishable. However, developing homozygous wri1seeds were impaired in the incorporation of sucrose and glucose into triacylglycerols, but incorporated pyruvate and acetate at an increased rate. Because the activities of several glycolytic enzymes, in particular hexokinase and pyrophosphate-dependent phosphofructokinase, are reduced in developing homozygouswri1 seeds, it is suggested that WRI1 is involved in the developmental regulation of carbohydrate metabolism during seed filling.

We have used cDNA microarrays to examine changes in gene expression during Arabidopsis seed development and to compare wild-type and mutant wrinkled1 (wri1) seeds that have an 80% reduction in oil. Between 5 and 13 days after flowering, a period preceding and including the major accumulation of storage oils and proteins, ∼35% of the genes represented on the array changed at least twofold, but a larger fraction (65%) showed little or no change in expression. Genes whose expression changed most tended to be expressed more in seeds than in other tissues. Genes related to the biosynthesis of storage components showed several distinct temporal expression patterns. For example, a number of genes encoding core fatty acid synthesis enzymes displayed a bell-shaped pattern of expression between 5 and 13 days after flowering. By contrast, the expression of storage proteins, oleosins, and other known abscisic acid–regulated genes increased later and remained high. Genes for photosynthetic proteins followed a pattern very similar to that of fatty acid synthesis proteins, implicating a role in CO2 refixation and the supply of cofactors for oil synthesis. Expression profiles of key carbon transporters and glycolytic enzymes reflected shifts in flux from cytosolic to plastid metabolism. Despite major changes in metabolism between wri1 and wild-type seeds, <1% of genes differed by more than twofold, and most of these were involved in central lipid and carbohydrate metabolism. Thus, these data define in part the downstream responses to disruption of the WRI1 gene.

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.