Identification of transcriptional modules linked to the drought response of Brassica napus during seed development and their mitigation by early biotic stress.

In order to capture the drought impacts on seed quality acquisition in Brassica napus and its potential interaction with early biotic stress, seeds of the 'Express' genotype of oilseed rape were characterized from late embryogenesis to full maturity from plants submitted to reduced watering (WS) with or without pre-occurring inoculation by the telluric pathogen Plasmodiophora brassicae (Pb + WS or Pb, respectively), and compared to control conditions (C). Drought as a single constraint led to significantly lower accumulation of lipids, higher protein content and reduced longevity of the WS-treated seeds. In contrast, when water shortage was preceded by clubroot infection, these phenotypic differences were completely abolished despite the upregulation of the drought sensor RD20. A weighted gene co-expression network of seed development in oilseed rape was generated using 72 transcriptomes from developing seeds from the four treatments and identified 33 modules. Module 29 was highly enriched in heat shock proteins and chaperones that showed a stronger upregulation in Pb + WS compared to the WS condition, pointing to a possible priming effect by the early P. brassicae infection on seed quality acquisition. Module 13 was enriched with genes encoding 12S and 2S seed storage proteins, with the latter being strongly upregulated under WS conditions. Cis-element promotor enrichment identified PEI1/TZF6, FUS3 and bZIP68 as putative regulators significantly upregulated upon WS compared to Pb + WS. Our results provide a temporal co-expression atlas of seed development in oilseed rape and will serve as a resource to characterize the plant response towards combinations of biotic and abiotic stresses.

Recent progress in molecular genetics and omics-driven research in seed biology.

Elucidating the mechanisms that control seed development, metabolism, and physiology is a fundamental issue in biology. Michel Caboche had long been a catalyst for seed biology research in France up until his untimely passing away last year. To honour his memory, we have updated a review written under his coordination in 2010 entitled "Arabidopsis seed secrets unravelled after a decade of genetic and omics-driven research". This review encompassed different molecular aspects of seed development, reserve accumulation, dormancy and germination, that are studied in the lab created by M. Caboche. We have extended the scope of this review to highlight original experimental approaches implemented in the field over the past decade such as omics approaches aimed at investigating the control of gene expression, protein modifications, primary and specialized metabolites at the tissue or even cellular level, as well as seed biodiversity and the impact of the environment on seed quality.

L'élucidation des mécanismes qui contrôlent le développement, le métabolisme et la physiologie des graines est une question fondamentale en biologie. Michel Caboche a longtemps été un catalyseur de la recherche en biologie des graines en France jusqu'à son décès prématuré l'année dernière. Pour honorer sa mémoire, nous avons mis à jour une revue écrite sous sa coordination en 2010 intitulée « Arabidopsis seed secrets unravelled after a decade of genetic and omics-driven research ¼. Cette revue englobait différents aspects moléculaires du développement des graines, de l'accumulation des réserves, de la dormance et de la germination, qui sont étudiés dans le laboratoire créé par M. Caboche. Nous avons étendu la portée de cette revue pour mettre en évidence des approches expérimentales originales mises en œuvre dans le domaine au cours de la dernière décennie, telles que les approches omiques visant à étudier le contrôle de l'expression des gènes, les modifications des protéines, les métabolites primaires et spécialisés au niveau des tissus ou même des cellules, tout en tenant compte de la biodiversité des graines et de l'impact de l'environnement sur leur qualité.

Plant monounsaturated fatty acids: Diversity, biosynthesis, functions and uses.

Monounsaturated fatty acids are straight-chain aliphatic monocarboxylic acids comprising a unique carbon­carbon double bond, also termed unsaturation. More than 50 distinct molecular structures have been described in the plant kingdom, and more remain to be discovered. The evolution of land plants has apparently resulted in the convergent evolution of non-homologous enzymes catalyzing the dehydrogenation of saturated acyl chain substrates in a chemo-, regio- and stereoselective manner. Contrasted enzymatic characteristics and different subcellular localizations of these desaturases account for the diversity of existing fatty acid structures. Interestingly, the location and geometrical configuration of the unsaturation confer specific characteristics to these molecules found in a variety of membrane, storage, and surface lipids. An ongoing research effort aimed at exploring the links existing between fatty acid structures and their biological functions has already unraveled the importance of several monounsaturated fatty acids in various physiological and developmental contexts. What is more, the monounsaturated acyl chains found in the oils of seeds and fruits are widely and increasingly used in the food and chemical industries due to the physicochemical properties inherent in their structures. Breeders and plant biotechnologists therefore develop new crops with high monounsaturated contents for various agro-industrial purposes.

Acyl-Acyl Carrier Protein Desaturases and Plant Biotic Interactions.

Interactions between land plants and other organisms such as pathogens, pollinators, or symbionts usually involve a variety of specialized effectors participating in complex cross-talks between organisms. Fatty acids and their lipid derivatives play important roles in these biological interactions. While the transcriptional regulation of genes encoding acyl-acyl carrier protein (ACP) desaturases appears to be largely responsive to biotic stress, the different monounsaturated fatty acids produced by these enzymes were shown to take active part in plant biotic interactions and were assigned with specific functions intrinsically linked to the position of the carbon-carbon double bond within their acyl chain. For example, oleic acid, an omega-9 monounsaturated fatty acid produced by Δ9-stearoyl-ACP desaturases, participates in signal transduction pathways affecting plant immunity against pathogen infection. Myristoleic acid, an omega-5 monounsaturated fatty acid produced by Δ9-myristoyl-ACP desaturases, serves as a precursor for the biosynthesis of omega-5 anacardic acids that are active biocides against pests. Finally, different types of monounsaturated fatty acids synthesized in the labellum of orchids are used for the production of a variety of alkenes participating in the chemistry of sexual deception, hence favoring plant pollination by hymenopterans.

Molecular Control of Oil Metabolism in the Endosperm of Seeds.

In angiosperm seeds, the endosperm develops to varying degrees and accumulates different types of storage compounds remobilized by the seedling during early post-germinative growth. Whereas the molecular mechanisms controlling the metabolism of starch and seed-storage proteins in the endosperm of cereal grains are relatively well characterized, the regulation of oil metabolism in the endosperm of developing and germinating oilseeds has received particular attention only more recently, thanks to the emergence and continuous improvement of analytical techniques allowing the evaluation, within a spatial context, of gene activity on one side, and lipid metabolism on the other side. These studies represent a fundamental step toward the elucidation of the molecular mechanisms governing oil metabolism in this particular tissue. In particular, they highlight the importance of endosperm-specific transcriptional controls for determining original oil compositions usually observed in this tissue. In the light of this research, the biological functions of oils stored in the endosperm of seeds then appear to be more diverse than simply constituting a source of carbon made available for the germinating seedling.

Docking of acetyl-CoA carboxylase to the plastid envelope membrane attenuates fatty acid production in plants.

In plants, light-dependent activation of de novo fatty acid synthesis (FAS) is partially mediated by acetyl-CoA carboxylase (ACCase), the first committed step for this pathway. However, it is not fully understood how plants control light-dependent FAS regulation to meet the cellular demand for acyl chains. We report here the identification of a gene family encoding for three small plastidial proteins of the envelope membrane that interact with the α-carboxyltransferase (α-CT) subunit of ACCase and participate in an original mechanism restraining FAS in the light. Light enhances the interaction between carboxyltransferase interactors (CTIs) and α-CT, which in turn attenuates carbon flux into FAS. Knockouts for CTI exhibit higher rates of FAS and marked increase in absolute triacylglycerol levels in leaves, more than 4-fold higher than in wild-type plants. Furthermore, WRINKLED1, a master transcriptional regulator of FAS, positively regulates CTI1 expression by direct binding to its promoter. This study reveals that in addition to light-dependent activation, "envelope docking" of ACCase permits fine-tuning of fatty acid supply during the plant life cycle.

Differential Activation of Partially Redundant Δ9 Stearoyl-ACP Desaturase Genes Is Critical for Omega-9 Monounsaturated Fatty Acid Biosynthesis During Seed Development in Arabidopsis.

The spatiotemporal pattern of deposition, final amount, and relative abundance of oleic acid (cis-ω-9 C18:1) and its derivatives in the different lipid fractions of the seed of Arabidopsis (Arabidopsis thaliana) indicates that omega-9 monoenes are synthesized at high rates in this organ. Accordingly, we observed that four Δ9 stearoyl-ACP desaturase (SAD)-coding genes (FATTY ACID BIOSYNTHESIS2 [FAB2], ACYL-ACYL CARRIER PROTEIN5 [AAD5], AAD1, and AAD6) are transcriptionally induced in seeds. We established that the three most highly expressed ones are directly activated by the WRINKLED1 transcription factor. We characterized a collection of 30 simple, double, triple, and quadruple mutants affected in SAD-coding genes and thereby revealed the functions of these desaturases throughout seed development. Production of oleic acid by FAB2 and AAD5 appears to be critical at the onset of embryo morphogenesis. Double homozygous plants from crossing fab2 and aad5 could never be obtained, and further investigations revealed that the double mutation results in the arrest of embryo development before the globular stage. During later stages of seed development, these two SADs, together with AAD1, participate in the elaboration of the embryonic cuticle, a barrier essential for embryo-endosperm separation during the phase of invasive embryo growth through the endosperm. This study also demonstrates that the four desaturases redundantly contribute to storage lipid production during the maturation phase.

AtMYB92 enhances fatty acid synthesis and suberin deposition in leaves of Nicotiana benthamiana.

Acyl lipids are important constituents of the plant cell. Depending on the cell type, requirements in acyl lipids vary greatly, implying a tight regulation of fatty acid and lipid metabolism. The discovery of the WRINKLED1 (WRI1) transcription factors, members of the AP2-EREBP (APETALA2-ethylene-responsive element binding protein) family, has emphasized the importance of transcriptional regulation for adapting the rate of acyl chain production to cell requirements. Here, we describe the identification of another activator of the fatty acid biosynthetic pathway, the Arabidopsis MYB92 transcription factor. This MYB and all the members of the subgroups S10 and S24 of MYB transcription factors can directly activate the promoter of BCCP2 that encodes a component of the fatty acid biosynthetic pathway. Two adjacent MYB cis-regulatory elements are essential for the binding and activation of the BCCP2 promoter by MYB92. Overexpression of MYB92 or WRI1 in Nicotiana benthamiana induces the expression of fatty acid biosynthetic genes but results in the accumulation of different types of acyl lipids. In the presence of WRI1, triacylglycerol biosynthetic enzymes coded by constitutively expressed genes efficiently channel the excess fatty acids toward reserve lipid accumulation. By contrast, MYB92 activates both fatty acid and suberin biosynthetic genes; hence, the remarkable increase in suberin monomers measured in leaves expressing MYB92. These results provide additional insight into the molecular mechanisms that control the biosynthesis of an important cell wall-associated acylglycerol polymer playing critical roles in plants.

Arabidopsis thaliana DGAT3 is a [2Fe-2S] protein involved in TAG biosynthesis.

Acyl-CoA:diacylglycerol acyltransferases 3 (DGAT3) are described as plant cytosolic enzymes synthesizing triacylglycerol. Their protein sequences exhibit a thioredoxin-like ferredoxin domain typical of a class of ferredoxins harboring a [2Fe-2S] cluster. The Arabidopsis thaliana DGAT3 (AtDGAT3; At1g48300) protein is detected in germinating seeds. The recombinant purified protein produced from Escherichia coli, although very unstable, exhibits DGAT activity in vitro. A shorter protein version devoid of its N-terminal putative chloroplast transit peptide, Δ46AtDGAT3, was more stable in vitro, allowing biochemical and spectroscopic characterization. The results obtained demonstrate the presence of a [2Fe-2S] cluster in the protein. To date, AtDGAT3 is the first metalloprotein described as a DGAT.

Seeds as oil factories.

KEY MESSAGE: Studying seed oil metabolism. The seeds of higher plants represent valuable factories capable of converting photosynthetically derived sugars into a variety of storage compounds, including oils. Oils are the most energy-dense plant reserves and fatty acids composing these oils represent an excellent nutritional source. They supply humans with much of the calories and essential fatty acids required in their diet. These oils are then increasingly being utilized as renewable alternatives to petroleum for the chemical industry and for biofuels. Plant oils therefore represent a highly valuable agricultural commodity, the demand for which is increasing rapidly. Knowledge regarding seed oil production is extensively exploited in the frame of breeding programs and approaches of metabolic engineering for oilseed crop improvement. Complementary aspects of this research include (1) the study of carbon metabolism responsible for the conversion of photosynthetically derived sugars into precursors for fatty acid biosynthesis, (2) the identification and characterization of the enzymatic actors allowing the production of the wide set of fatty acid structures found in seed oils, and (3) the investigation of the complex biosynthetic pathways leading to the production of storage lipids (waxes, triacylglycerols). In this review, we outline the most recent developments in our understanding of the underlying biochemical and molecular mechanisms of seed oil production, focusing on fatty acids and oils that can have a significant impact on the emerging bioeconomy.

Overexpression of MYB115, AAD2, or AAD3 in Arabidopsis thaliana seeds yields contrasting omega-7 contents.

Omega-7 monoenoic fatty acids (ω-7 FAs) are increasingly exploited both for their positive effects on health and for their industrial potential. Some plant species produce fruits or seeds with high amounts of ω-7 FAs. However, the low yields and poor agronomic properties of these plants preclude their commercial use. As an alternative, the metabolic engineering of oilseed crops for sustainable ω-7 FA production has been proposed. Two palmitoyl-ACP desaturases (PADs) catalyzing ω-7 FA biosynthesis were recently identified and characterized in Arabidopsis thaliana, together with MYB115 and MYB118, two transcription factors that positively control the expression of the corresponding PAD genes. In the present research, we examine the biotechnological potential of these new actors of ω-7 metabolism for the metabolic engineering of plant-based production of ω-7 FAs. We placed the PAD and MYB115 coding sequences under the control of a promoter strongly induced in seeds and evaluated these different constructs in A. thaliana. Seeds were obtained that exhibit ω-7 FA contents ranging from 10 to >50% of the total FAs, and these major compositional changes have no detrimental effect on seed germination.

Transcriptional Activation of Two Delta-9 Palmitoyl-ACP Desaturase Genes by MYB115 and MYB118 Is Critical for Biosynthesis of Omega-7 Monounsaturated Fatty Acids in the Endosperm of Arabidopsis Seeds.

In angiosperms, double fertilization of the embryo sac initiates the development of the embryo and the endosperm. In Arabidopsis thaliana, an exalbuminous species, the endosperm is reduced to one cell layer during seed maturation and reserves such as oil are massively deposited in the enlarging embryo. Here, we consider the strikingly different fatty acid (FA) compositions of the oils stored in the two zygotic tissues. Endosperm oil is enriched in ω-7 monounsaturated FAs, that represent more than 20 mol% of total FAs, whereas these molecular species are 10-fold less abundant in the embryo. Two closely related transcription factors, MYB118 and MYB115, are transcriptionally induced at the onset of the maturation phase in the endosperm and share a set of transcriptional targets. Interestingly, the endosperm oil of myb115 myb118 double mutants lacks ω-7 FAs. The identification of two Δ9 palmitoyl-ACP desaturases responsible for ω-7 FA biosynthesis, which are activated by MYB115 and MYB118 in the endosperm, allows us to propose a model for the transcriptional control of oil FA composition in this tissue. In addition, an initial characterization of the structure-function relationship for these desaturases reveals that their particular substrate specificity is conferred by amino acid residues lining their substrate pocket that distinguish them from the archetype Δ9 stearoyl-ACP desaturase.

Deciphering and modifying LAFL transcriptional regulatory network in seed for improving yield and quality of storage compounds.

Increasing yield and quality of seed storage compounds in a sustainable way is a key challenge for our societies. Genome-wide analyses conducted in both monocot and dicot angiosperms emphasized drastic transcriptional switches that occur during seed development. In Arabidopsis thaliana, a reference species, genetic and molecular analyses have demonstrated the key role of LAFL (LEC1, ABI3, FUS3, and LEC2) transcription factors (TFs), in controlling gene expression programs essential to accomplish seed maturation and the accumulation of storage compounds. Here, we summarize recent progress obtained in the characterization of these LAFL proteins, their regulation, partners and target genes. Moreover, we illustrate how these evolutionary conserved TFs can be used to engineer new crops with altered seed compositions and point out the current limitations. Last, we discuss about the interest of investigating further the environmental and epigenetic regulation of this network for the coming years.

Deciphering the Molecular Mechanisms Underpinning the Transcriptional Control of Gene Expression by Master Transcriptional Regulators in Arabidopsis Seed.

In Arabidopsis (Arabidopsis thaliana), transcriptional control of seed maturation involves three related regulators with a B3 domain, namely LEAFY COTYLEDON2 (LEC2), ABSCISIC ACID INSENSITIVE3 (ABI3), and FUSCA3 (ABI3/FUS3/LEC2 [AFLs]). Although genetic analyses have demonstrated partially overlapping functions of these regulators, the underlying molecular mechanisms remained elusive. The results presented here confirmed that the three proteins bind RY DNA elements (with a 5'-CATG-3' core sequence) but with different specificities for flanking nucleotides. In planta as in the moss Physcomitrella patens protoplasts, the presence of RY-like (RYL) elements is necessary but not sufficient for the regulation of the OLEOSIN1 (OLE1) promoter by the B3 AFLs. G box-like domains, located in the vicinity of the RYL elements, also are required for proper activation of the promoter, suggesting that several proteins are involved. Consistent with this idea, LEC2 and ABI3 showed synergistic effects on the activation of the OLE1 promoter. What is more, LEC1 (a homolog of the NF-YB subunit of the CCAAT-binding complex) further enhanced the activation of this target promoter in the presence of LEC2 and ABI3. Finally, recombinant LEC1 and LEC2 proteins produced in Arabidopsis protoplasts could form a ternary complex with NF-YC2 in vitro, providing a molecular explanation for their functional interactions. Taken together, these results allow us to propose a molecular model for the transcriptional regulation of seed genes by the L-AFL proteins, based on the formation of regulatory multiprotein complexes between NF-YBs, which carry a specific aspartate-55 residue, and B3 transcription factors.

New insights on the organization and regulation of the fatty acid biosynthetic network in the model higher plant Arabidopsis thaliana.

In the plastids of plant cells, fatty acid (FA) production is a central biosynthetic process. It provides acyl chains for the formation of a variety of acyl lipids fulfilling different biological functions ranging from membrane synthesis to signaling or carbon and energy storage. The biochemical pathway leading to the synthesis of FA has been described for a long time. Over the last 15 years, and after the genome of the model higher plant Arabidopsis thaliana has been sequenced, the scientific community has deployed approaches of functional genomics to identify the actors comprising this pathway. One of the puzzling aspects of the emerging molecular biology of FA synthesis resided in the occurrence of multigene families encoding most enzymes of the pathway. Studies carried out to investigate these families led to the conclusion that most members have acquired non-redundant roles in planta. This is usually the consequence of divergent expression patterns of these isogenes and/or of different substrate specificities of the isoforms they encode. Nevertheless, much remains to be elucidated regarding the molecular bases underpinning these specificities. Protein biochemistry together with emerging quantitative proteomic technologies have then led to a better understanding of the structure of the network, which is composed of multiprotein complexes organized within the stromal compartment of plastids: whereas growing evidence suggests that the early steps of the pathway might be associated to the inner envelope membrane, several late enzymes might be localized next to the thylakoids. The question of the existence of a large integrated protein assembly channeling substrates through the whole pathway that would span the stroma remains uncertain. Finally, recent discoveries regarding the post-translational regulation of the pathway open new research horizons and may guide the development of relevant biotechnological strategies aimed at monitoring FA production in plant systems.

MYB118 represses endosperm maturation in seeds of Arabidopsis.

In the exalbuminous species Arabidopsis thaliana, seed maturation is accompanied by the deposition of oil and storage proteins and the reduction of the endosperm to one cell layer. Here, we consider reserve partitioning between embryo and endosperm compartments. The pattern of deposition, final amount, and composition of these reserves differ between the two compartments, with the embryo representing the principal storage tissue in mature seeds. Complex regulatory mechanisms are known to prevent activation of maturation-related programs during embryo morphogenesis and, later, during vegetative growth. Here, we describe a regulator that represses the expression of maturation-related genes during maturation within the endosperm. MYB118 is transcriptionally induced in the maturing endosperm, and seeds of myb118 mutants exhibit an endosperm-specific derepression of maturation-related genes associated with a partial relocation of storage compounds from the embryo to the endosperm. Moreover, MYB118 activates endosperm-induced genes through the recognition of TAACGG elements. These results demonstrate that the differential partitioning of reserves between the embryo and endosperm in exalbuminous Arabidopsis seeds does not only result from developmental programs that establish the embryo as the preponderant tissue within seeds. This differential partitioning is also regulated by MYB118, which regulates the biosynthesis of reserves at the spatial level during maturation.

Function and localization of the Arabidopsis thaliana diacylglycerol acyltransferase DGAT2 expressed in yeast.

Diacylglycerol acyltransferases (DGATs) catalyze the final and only committed step of triacylglycerol synthesis. DGAT activity is rate limiting for triacylglycerol accumulation in mammals, plants and microbes. DGATs belong to three different evolutionary classes. In Arabidopsis thaliana, DGAT1, encoded by At2g19450, is the major DGAT enzyme involved in triacylglycerol accumulation in seeds. Until recently, the function of DGAT2 (At3g51520) has remained elusive. Previous attempts to characterize its enzymatic function by heterologous expression in yeast were unsuccessful. In the present report we demonstrate that expression of a codon-optimized version of the DGAT2 gene is able to restore neutral lipid accumulation in the Saccharomyces cerevisiae mutant strain (H1246), which is defective in triacylglycerol biosynthesis. Heterologous expression of codon-optimized DGAT2 and DGAT1 induced the biogenesis of subcellular lipid droplets containing triacylglycerols and squalene. Both DGAT proteins were found to be associated with these lipid droplets. The fatty acid composition was affected by the nature of the acyltransferase expressed. DGAT2 preferentially incorporated C16:1 fatty acids whereas DGAT1 displayed preference for C16:0, strongly suggesting that these enzymes have contrasting substrate specificities.

Specialization of oleosins in oil body dynamics during seed development in Arabidopsis seeds.

Oil bodies (OBs) are seed-specific lipid storage organelles that allow the accumulation of neutral lipids that sustain plantlet development after the onset of germination. OBs are covered with specific proteins embedded in a single layer of phospholipids. Using fluorescent dyes and confocal microscopy, we monitored the dynamics of OBs in living Arabidopsis (Arabidopsis thaliana) embryos at different stages of development. Analyses were carried out with different genotypes: the wild type and three mutants affected in the accumulation of various oleosins (OLE1, OLE2, and OLE4), three major OB proteins. Image acquisition was followed by a detailed statistical analysis of OB size and distribution during seed development in the four dimensions (x, y, z, and t). Our results indicate that OB size increases sharply during seed maturation, in part by OB fusion, and then decreases until the end of the maturation process. In single, double, and triple mutant backgrounds, the size and spatial distribution of OBs are modified, affecting in turn the total lipid content, which suggests that the oleosins studied have specific functions in the dynamics of lipid accumulation.

Acyl-lipid metabolism.

Acyl lipids in Arabidopsis and all other plants have a myriad of diverse functions. These include providing the core diffusion barrier of the membranes that separates cells and subcellular organelles. This function alone involves more than 10 membrane lipid classes, including the phospholipids, galactolipids, and sphingolipids, and within each class the variations in acyl chain composition expand the number of structures to several hundred possible molecular species. Acyl lipids in the form of triacylglycerol account for 35% of the weight of Arabidopsis seeds and represent their major form of carbon and energy storage. A layer of cutin and cuticular waxes that restricts the loss of water and provides protection from invasions by pathogens and other stresses covers the entire aerial surface of Arabidopsis. Similar functions are provided by suberin and its associated waxes that are localized in roots, seed coats, and abscission zones and are produced in response to wounding. This chapter focuses on the metabolic pathways that are associated with the biosynthesis and degradation of the acyl lipids mentioned above. These pathways, enzymes, and genes are also presented in detail in an associated website (ARALIP: http://aralip.plantbiology.msu.edu/). Protocols and methods used for analysis of Arabidopsis lipids are provided. Finally, a detailed summary of the composition of Arabidopsis lipids is provided in three figures and 15 tables.

WRINKLED transcription factors orchestrate tissue-specific regulation of fatty acid biosynthesis in Arabidopsis.

Acyl lipids are essential constituents of all cells, but acyl chain requirements vary greatly and depend on the cell type considered. This implies a tight regulation of fatty acid production so that supply fits demand. Isolation of the Arabidopsis thaliana WRINKLED1 (WRI1) transcription factor established the importance of transcriptional regulation for modulating the rate of acyl chain production. Here, we report the isolation of two additional regulators of the fatty acid biosynthetic pathway, WRI3 and WRI4, which are closely related to WRI1 and belong to the APETALA2-ethylene-responsive element binding protein family of transcription factors. These three WRIs define a family of regulators capable of triggering sustained rates of acyl chain synthesis. However, expression patterns of the three WRIs differ markedly. Whereas only WRI1 activates fatty acid biosynthesis in seeds for triacylglycerol production, the three WRIs are required in floral tissues to provide acyl chains for cutin biosynthesis and prevent adherence of these developing organs and subsequent semisterility. The targets of these WRIs encode enzymes providing precursors (acyl chain and glycerol backbones) for various lipid biosynthetic pathways, but not the subsequent lipid-assembling enzymes. These results provide insights into the developmental regulation of fatty acid production in plants.