Transcriptomes across fertilization and seed development in the water lily Nymphaea thermarum (Nymphaeales): evidence for epigenetic patterning during reproduction.

KEY MESSAGE: The first record of gene expression during seed development within the Nymphaeales provides evidence for a variety of biological processes, including dynamic epigenetic patterning during sexual reproduction in the water lily Nymphaea thermarum. Studies of gene expression during seed development have been performed for a growing collection of species from a phylogenetically broad sampling of flowering plants (angiosperms). However, angiosperm lineages whose origins predate the divergence of monocots and eudicots have been largely overlooked. In order to provide a new resource for understanding the early evolution of seed development in flowering plants, we sequenced transcriptomes of whole ovules and seeds from three key stages of reproductive development in the waterlily Nymphaea thermarum, an experimentally tractable member of the Nymphaeales. We first explore patterns of gene expression, beginning with mature ovules and continuing through fertilization into early- and mid-stages of seed development. We find patterns of gene expression that corroborate histological/morphological observations of seed development in this species, such as expression of genes involved in starch synthesis and transcription factors that have been associated with embryo and endosperm development in other species. We also find evidence for processes that were previously not known to be occurring during seed development in this species, such as epigenetic modification. We then examine the expression of genes associated with patterning DNA and histone methylation-processes that are essential for seed development in distantly related and structurally diverse monocots and eudicots. Around 89% of transcripts putatively homologous to DNA and histone methylation modifiers are expressed during seed development in N. thermarum, including homologs of genes known to pattern imprinting-related epigenetic modifications. Our results suggest that dynamic epigenetic patterning is a deeply conserved aspect of angiosperm seed development.

Maternal-filial transfer structures in endosperm: A nexus of nutritional dynamics and seed development.

Although the ultimate purpose of a seed is the successful establishment of the next generation, seed development involves more than embryo growth. In angiosperms, seed development requires the intimate coordination of three distinct entities - maternal tissue and two offspring, embryo and embryo-nourishing endosperm. Although seeds are cornerstones of many terrestrial ecosystems and human diets, we are only beginning to understand the interactions among seed tissues and the molecular processes and genes that determine them. Recent studies of gene expression and function in distantly related angiosperms, combined with over 100 years of embryological research, have repeatedly highlighted the endosperm associated with maternal-filial boundaries as a central point in seed developmental dynamics. In this review, we highlight evidence that links this zone with nutritional dynamics, developmental signaling, and imprinted gene expression. We suggest that the underappreciated diversity of this specialized endosperm across angiosperms deserves further study from developmental, molecular, and genetic perspectives.

Transcriptional and imprinting complexity in Arabidopsis seeds at single-nucleus resolution.

Seeds are a key life cycle stage for many plants. Seeds are also the basis of agriculture and the primary source of calories consumed by humans1. Here, we employ single-nucleus RNA-sequencing to generate a transcriptional atlas of developing Arabidopsis thaliana seeds, with a focus on endosperm. Endosperm, the primary site of gene imprinting in flowering plants, mediates the relationship between the maternal parent and the embryo2. We identify transcriptionally uncharacterized nuclei types in the chalazal endosperm, which interfaces with maternal tissue for nutrient unloading3,4. We demonstrate that the extent of parental bias of maternally expressed imprinted genes varies with cell-cycle phase, and that imprinting of paternally expressed imprinted genes is strongest in chalazal endosperm. Thus, imprinting is spatially and temporally heterogeneous. Increased paternal expression in the chalazal region suggests that parental conflict, which is proposed to drive imprinting evolution, is fiercest at the boundary between filial and maternal tissues.

Water lily (Nymphaea thermarum) genome reveals variable genomic signatures of ancient vascular cambium losses.

For more than 225 million y, all seed plants were woody trees, shrubs, or vines. Shortly after the origin of angiosperms ∼140 million y ago (MYA), the Nymphaeales (water lilies) became one of the first lineages to deviate from their ancestral, woody habit by losing the vascular cambium, the meristematic population of cells that produces secondary xylem (wood) and phloem. Many of the genes and gene families that regulate differentiation of secondary tissues also regulate the differentiation of primary xylem and phloem, which are produced by apical meristems and retained in nearly all seed plants. Here, we sequenced and assembled a draft genome of the water lily Nymphaea thermarum, an emerging system for the study of early flowering plant evolution, and compared it to genomes from other cambium-bearing and cambium-less lineages (e.g., monocots and Nelumbo). This revealed lineage-specific patterns of gene loss and divergence. Nymphaea is characterized by a significant contraction of the HD-ZIP III transcription factors, specifically loss of REVOLUTA, which influences cambial activity in other angiosperms. We also found the Nymphaea and monocot copies of cambium-associated CLE signaling peptides display unique substitutions at otherwise highly conserved amino acids. Nelumbo displays no obvious divergence in cambium-associated genes. The divergent genomic signatures of convergent loss of vascular cambium reveals that even pleiotropic genes can exhibit unique divergence patterns in association with independent events of trait loss. Our results shed light on the evolution of herbaceousness-one of the key biological innovations associated with the earliest phases of angiosperm evolution.

Evidence for parent-of-origin effects and interparental conflict in seeds of an ancient flowering plant lineage.

Theoretical and empirical studies have long connected the evolutionary innovation of endosperm, a genetically biparental product of a double fertilization process unique to flowering plants (angiosperms), to conflicting parental interests over offspring provisioning. Yet, none of these studies examined interparental conflict in representatives of any of the most ancient angiosperm lineages. We performed reciprocal interploidy crosses in the water lily Nymphaea thermarum, a member of one of the most ancient angiosperm lineages, Nymphaeales. We find that an excess of paternal genomes is associated with an increase in endosperm growth. By contrast, maternal ploidy negatively influences development or growth of all seed components, regardless of paternal genome dosage. Most relevant to the conflict over distribution of maternal resources, however, is that growth of the perisperm (seed storage tissue derived from the maternal sporophyte, found in all Nymphaeales) is unaffected by paternal genome dosage-ensuring maternal control of maternal resources. We conclude that the evolutionary transfer of embryo-nourishing function from a genetically biparental endosperm to a genetically maternal perisperm can be viewed as an effective maternal strategy to recapture control of resource distribution among progeny, and thus that interparental conflict has influenced the evolution of seed development in this ancient angiosperm lineage.

Floral biology and ovule and seed ontogeny of Nymphaea thermarum, a water lily at the brink of extinction with potential as a model system for basal angiosperms.

BACKGROUND AND AIMS: Nymphaea thermarum is a member of the Nymphaeales, of one of the most ancient lineages of flowering plants. This species was only recently described and then declared extinct in the wild, so little is known about its reproductive biology. In general, the complete ontogeny of ovules and seeds is not well documented among species of Nymphaea and has never been studied in the subgenus Brachyceras, the clade to which N. thermarum belongs. METHODS: Flowers and fruits were processed for brightfield, epifluorescence and confocal microscopy. Flower morphology, with emphasis on the timing of male and female functions, was correlated with key developmental stages of the ovule and the female gametophyte. Development of the seed tissues and dynamics of polysaccharide reserves in the endosperm, perisperm and embryo were examined. KEY RESULTS: Pollen release in N. thermarum starts before the flower opens. Cell walls of the micropylar nucellus show layering of callose and cellulose in a manner reminiscent of transfer cell wall patterning. Endosperm development is ab initio cellular, with micropylar and chalazal domains that embark on distinct developmental trajectories. The surrounding maternal perisperm occupies the majority of seed volume and accumulates starch centrifugally. In mature seeds, a minute but fully developed embryo is surrounded by a single, persistent layer of endosperm. CONCLUSIONS: Early male and female function indicate that N. thermarum is predisposed towards self-pollination, a phenomenon that is likely to have evolved multiple times within Nymphaea. While formation of distinct micropylar and chalazal developmental domains in the endosperm, along with a copious perisperm, characterize the seeds of most members of the Nymphaeales, seed ontogenies vary between and among the constituent families. Floral biology, life history traits and small genome size make N. thermarum uniquely promising as an early-diverging angiosperm model system for genetic and molecular studies.