ABSTRACT
The nucleus is composed of functionally distinct membraneless compartments that undergo phase separation (PS). However, whether different subnuclear compartments are connected remains elusive. We identified a type of nuclear body with PS features composed of BAZ2A that associates with active chromatin. BAZ2A bodies depend on RNA transcription and BAZ2A non-disordered RNA-binding TAM domain. Although BAZ2A and H3K27me3 occupancies anticorrelate in the linear genome, in the nuclear space, BAZ2A bodies contact H3K27me3 bodies. BAZ2A-body disruption promotes BAZ2A invasion into H3K27me3 domains, causing H3K27me3-body loss and gene upregulation. Weak BAZ2A-RNA interactions, such as with nascent transcripts, promote BAZ2A bodies, whereas the strong binder long non-coding RNA (lncRNA) Malat1 impairs them while mediating BAZ2A association to chromatin at nuclear speckles. In addition to unraveling a direct connection between nuclear active and repressive compartments through PS mechanisms, the results also showed that the strength of RNA-protein interactions regulates this process, contributing to nuclear organization and the regulation of chromatin and gene expression.
Subject(s)
Chromatin , Histones , RNA, Long Noncoding , Chromatin/metabolism , Chromatin/genetics , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Humans , Histones/metabolism , Histones/genetics , Cell Nucleus/metabolism , Cell Nucleus/genetics , RNA-Binding Proteins/metabolism , RNA-Binding Proteins/genetics , HeLa Cells , Transcription, Genetic , RNA/metabolism , RNA/genetics , Animals , Gene Expression RegulationABSTRACT
Plant protoplasts provide starting material for of inducing pluripotent cell masses that are competent for tissue regeneration in vitro, analogous to animal induced pluripotent stem cells (iPSCs). Dedifferentiation is associated with large-scale chromatin reorganisation and massive transcriptome reprogramming, characterised by stochastic gene expression. How this cellular variability reflects on chromatin organisation in individual cells and what factors influence chromatin transitions during culturing are largely unknown. Here, we used high-throughput imaging and a custom supervised image analysis protocol extracting over 100 chromatin features of cultured protoplasts. The analysis revealed rapid, multiscale dynamics of chromatin patterns with a trajectory that strongly depended on nutrient availability. Decreased abundance in H1 (linker histones) is hallmark of chromatin transitions. We measured a high heterogeneity of chromatin patterns indicating intrinsic entropy as a hallmark of the initial cultures. We further measured an entropy decline over time, and an antagonistic influence by external and intrinsic factors, such as phytohormones and epigenetic modifiers, respectively. Collectively, our study benchmarks an approach to understand the variability and evolution of chromatin patterns underlying plant cell reprogramming in vitro.
Subject(s)
Chromatin , Entropy , Induced Pluripotent Stem Cells , Chromatin/metabolism , Chromatin/genetics , Induced Pluripotent Stem Cells/metabolism , Induced Pluripotent Stem Cells/cytology , Protoplasts/metabolism , Cellular Reprogramming/genetics , Histones/metabolism , Histones/genetics , Plant Cells/metabolism , Epigenesis, GeneticABSTRACT
The nucleoskeleton forms a filamentous meshwork under the nuclear envelope and contributes to the regulation of nuclear shape and gene expression. To understand how the Arabidopsis (Arabidopsis thaliana) nucleoskeleton physically connects to the nuclear periphery in plants, we investigated the Arabidopsis nucleoskeleton protein KAKU4 and sought for functional regions responsible for its localization at the nuclear periphery. We identified 3 conserved peptide motifs within the N-terminal region of KAKU4, which are required for intermolecular interactions of KAKU4 with itself, interaction with the nucleoskeleton protein CROWDED NUCLEI (CRWN), localization at the nuclear periphery, and nuclear elongation in differentiated tissues. Unexpectedly, we find these motifs to be present also in NUP82 and NUP136, 2 plant-specific nucleoporins from the nuclear pore basket. We further show that NUP82, NUP136, and KAKU4 have a common evolutionary history predating nonvascular land plants with KAKU4 mainly localizing outside the nuclear pore suggesting its divergence from an ancient nucleoporin into a new nucleoskeleton component. Finally, we demonstrate that both NUP82 and NUP136, through their shared N-terminal motifs, interact with CRWN and KAKU4 proteins revealing the existence of a physical continuum between the nuclear pore and the nucleoskeleton in plants.
Subject(s)
Arabidopsis Proteins , Arabidopsis , Nuclear Pore/genetics , Nuclear Pore/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Amino Acid Motifs , Nuclear Envelope/genetics , Nuclear Envelope/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Nuclear Pore Complex Proteins/genetics , Nuclear Pore Complex Proteins/metabolism , Nuclear Matrix/metabolismABSTRACT
Defining the contributions and interactions of paternal and maternal genomes during embryo development is critical to understand the fundamental processes involved in hybrid vigor, hybrid sterility, and reproductive isolation. To determine the parental contributions and their regulation during Arabidopsis embryogenesis, we combined deep-sequencing-based RNA profiling and genetic analyses. At the 2-4 cell stage there is a strong, genome-wide dominance of maternal transcripts, although transcripts are contributed by both parental genomes. At the globular stage the relative paternal contribution is higher, largely due to a gradual activation of the paternal genome. We identified two antagonistic maternal pathways that control these parental contributions. Paternal alleles are initially downregulated by the chromatin siRNA pathway, linked to DNA and histone methylation, whereas transcriptional activation requires maternal activity of the histone chaperone complex CAF1. Our results define maternal epigenetic pathways controlling the parental contributions in plant embryos, which are distinct from those regulating genomic imprinting.
Subject(s)
Arabidopsis/embryology , Arabidopsis/genetics , Epigenomics , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Arabidopsis Proteins/metabolism , Gene Expression Profiling , Genome, Plant , Histone-Lysine N-Methyltransferase/metabolism , Ovule/metabolism , RNA Splicing Factors , RNA, Small Interfering/metabolism , Seeds/genetics , Transcriptional ActivationABSTRACT
Rabl organization is a type of interphase chromosome arrangement with centromeres and telomeres clustering at opposite nuclear poles. Here, we analyzed nuclear morphology and chromosome organization in cycling and endoreduplicated nuclei isolated from embryo and endosperm tissues of developing barley seeds. We show that endoreduplicated nuclei have an irregular shape, less sister chromatid cohesion at 5S rDNA loci, and a reduced amount of centromeric histone CENH3. While the chromosomes of the embryo and endosperm nuclei are initially organized in Rabl configuration, the centromeres and telomeres are intermingled within the nuclear space in the endoreduplicated nuclei with an increasing endoreduplication level. Such a loss of chromosome organization suggests that Rabl configuration is introduced and further reinforced by mitotic divisions in barley cell nuclei in a tissue- and seed age-dependent manner.
Subject(s)
Hordeum , Hordeum/genetics , Endosperm/genetics , Cell Nucleus/genetics , Histones/genetics , Centromere/geneticsABSTRACT
This Community Resource paper introduces the range of materials developed by the INDEPTH (Impact of Nuclear Domains on Gene Expression and Plant Traits) COST Action made available through the INDEPTH Academy. Recent rapid growth in understanding of the significance of epigenetic controls in plant and crop science has led to a need for shared, high-quality resources, standardization of protocols, and repositories for open access data. The INDEPTH Academy provides a range of masterclass tutorials, standardized protocols, and teaching webinars, together with a rapidly developing repository to support imaging and spatial analysis of the nucleus and deep learning for automated analysis. These resources were developed partly as a response to the COVID-19 pandemic, but also driven by needs and opportunities identified by the INDEPTH community of ~200 researchers in 80 laboratories from 32 countries. This community report outlines the resources produced and how they will be extended beyond the INDEPTH project, but also aims to encourage the wider community to engage with epigenetics and nuclear structure by accessing these resources.
Subject(s)
COVID-19 , Community Resources , Gene Expression , Humans , Pandemics , Plants/geneticsABSTRACT
The maintenance of genome integrity over cell divisions is critical for plant development and the correct transmission of genetic information to the progeny. A key factor involved in this process is the STRUCTURAL MAINTENANCE OF CHROMOSOME5 (SMC5) and SMC6 (SMC5/6) complex, related to the cohesin and condensin complexes that control sister chromatid alignment and chromosome condensation, respectively. Here, we characterize NON-SMC ELEMENT4 (NSE4) paralogs of the SMC5/6 complex in Arabidopsis (Arabidopsis thaliana). NSE4A is expressed in meristems and accumulates during DNA damage repair. Partial loss-of-function nse4a mutants are viable but hypersensitive to DNA damage induced by zebularine. In addition, nse4a mutants produce abnormal seeds, with noncellularized endosperm and embryos that maximally develop to the heart or torpedo stage. This phenotype resembles the defects in cohesin and condensin mutants and suggests a role for all three SMC complexes in differentiation during seed development. By contrast, NSE4B is expressed in only a few cell types, and loss-of-function mutants do not have any obvious abnormal phenotype. In summary, our study shows that the NSE4A subunit of the SMC5-SMC6 complex is essential for DNA damage repair in somatic tissues and plays a role in plant reproduction.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/embryology , Cell Cycle Proteins/metabolism , DNA Damage , DNA Repair , Protein Subunits/metabolism , Seeds/metabolism , Arabidopsis/genetics , Arabidopsis/immunology , Arabidopsis Proteins/genetics , Cell Cycle Proteins/genetics , DNA Damage/genetics , DNA Repair/genetics , Gene Duplication , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Genome, Plant , Ovule/genetics , Pollen/genetics , Protein Binding , Seeds/genetics , Transcriptome/genetics , Up-Regulation/geneticsABSTRACT
The precise location of chromatin domains within the cell nucleus has seen growing recognition in the past decade as an additional mechanism of controlling gene expression in both plants and animals (Dekker et al., 2017). Consequently, international efforts are devoted to understanding the organising principle of this organelle in plants, and notably the nature and the role of functional compartments on gene expression (Graumann et al., 2013; Sotelo-Silveira et al., 2018). The European cooperation 'Impact of Nuclear Domains on Gene Expression and Plant Traits' (INDEPTH) brings together molecular cell biologists, plant physiologists, bioinformaticians, image analysts and computer scientists. They aim to address the question of how nuclear architecture, chromatin organisation and gene expression are connected in plants, particularly in relation to traits of interest such as biomass, reproduction and resistance to pathogens (https://www.brookes.ac.uk/indepth/). The kick-off meeting of the INDEPTH consortium took place in Clermont-Ferrand, France, on 12-14th March 2018, where more than 80 researchers set the agenda for the coming four years of research and collaboration.
Subject(s)
Cell Nucleus/metabolism , Chromatin/metabolism , HumansABSTRACT
Embryonic development requires a correct balancing of maternal and paternal genetic information. This balance is mediated by genomic imprinting, an epigenetic mechanism that leads to parent-of-origin-dependent gene expression. The parental conflict (or kinship) theory proposes that imprinting can evolve due to a conflict between maternal and paternal alleles over resource allocation during seed development. One assumption of this theory is that paternal alleles can regulate seed growth; however, paternal effects on seed size are often very low or non-existent. We demonstrate that there is a pool of cryptic genetic variation in the paternal control of Arabidopsis thaliana seed development. Such cryptic variation can be exposed in seeds that maternally inherit a medea mutation, suggesting that MEA acts as a maternal buffer of paternal effects. Genetic mapping using recombinant inbred lines, and a novel method for the mapping of parent-of-origin effects using whole-genome sequencing of segregant bulks, indicate that there are at least six loci with small, paternal effects on seed development. Together, our analyses reveal the existence of a pool of hidden genetic variation on the paternal control of seed development that is likely shaped by parental conflict.
Subject(s)
Arabidopsis/genetics , Genetic Variation , Genomic Imprinting , Seeds/genetics , Alleles , Arabidopsis/growth & development , Gene Expression Regulation, Plant , Genome, Plant , Models, Genetic , Plant Development/genetics , Seeds/growth & developmentABSTRACT
The endosperm is at the center of successful seed formation in flowering plants. Being itself a product of fertilization, it is devoted to nourish the developing embryo and typically possesses a triploid genome consisting of two maternal and one paternal genome complement. Interestingly, endosperm development is controlled by epigenetic mechanisms conferring parent-of-origin-dependent effects that influence seed development. In the model plant Arabidopsis thaliana, we have previously described an endosperm-specific heterochromatin fraction, which increases with higher maternal, but not paternal, genome dosage. Here, we report a detailed analysis of chromosomal arrangement and association frequency in endosperm nuclei. We found that centromeric FISH signals in isolated nuclei show a planar alignment that may results from a semi-rigid, connective structure between chromosomes. Importantly, we found frequent pairwise association of centromeres, chromosomal segments, and entire arms of chromosomes in 3C endosperm nuclei. These associations deviate from random expectations predicted by numerical simulations. Therefore, we suggest a non-random chromosomal organization in the triploid nuclei of Arabidopsis endosperm. This contrasts with the prevailing random arrangement of chromosome territories in somatic nuclei. Based on observations on a series of nuclei with varying parental genome ratios, we propose a model where chromosomes associate pairwise involving one maternal and one paternal complement. The functional implications of this predicted chromosomal arrangement are discussed.
Subject(s)
Cell Nucleus/genetics , Chromosomes, Plant , Endosperm/genetics , Translocation, Genetic , Triploidy , Arabidopsis/genetics , Centromere , Gene Dosage , Genome, Plant , Seeds/geneticsABSTRACT
H1 (or linker) histones are basic nuclear proteins that possess an evolutionarily conserved nucleosome-binding globular domain, GH1. They perform critical functions in determining the accessibility of chromatin DNA to trans-acting factors. In most metazoan species studied so far, linker histones are highly heterogenous, with numerous nonallelic variants cooccurring in the same cells. The phylogenetic relationships among these variants as well as their structural and functional properties have been relatively well established. This contrasts markedly with the rather limited knowledge concerning the phylogeny and structural and functional roles of an unusually diverse group of GH1-containing proteins in plants. The dearth of information and the lack of a coherent phylogeny-based nomenclature of these proteins can lead to misunderstandings regarding their identity and possible relationships, thereby hampering plant chromatin research. Based on published data and our in silico and high-throughput analyses, we propose a systematization and coherent nomenclature of GH1-containing proteins of Arabidopsis (Arabidopsis thaliana [L.] Heynh) that will be useful for both the identification and structural and functional characterization of homologous proteins from other plant species.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , Histones/genetics , Phylogeny , Arabidopsis/metabolism , Arabidopsis Proteins/classification , Arabidopsis Proteins/metabolism , Binding Sites/genetics , Databases, Genetic , Databases, Protein , Histones/classification , Histones/metabolism , Plant Proteins/classification , Plant Proteins/genetics , Plant Proteins/metabolism , Plants/classification , Plants/genetics , Plants/metabolism , Species SpecificityABSTRACT
Sexual reproduction in flowering plants offers a number of remarkable aspects to developmental biologists. First, the spore mother cells - precursors of the plant reproductive lineage - are specified late in development, as opposed to precocious germline isolation during embryogenesis in most animals. Second, unlike in most animals where meiosis directly produces gametes, plant meiosis entails the differentiation of a multicellular, haploid gametophyte, within which gametic as well as non-gametic accessory cells are formed. These observations raise the question of the factors inducing and modus operandi of cell fate transitions that originate in floral tissues and gametophytes, respectively. Cell fate transitions in the reproductive lineage imply cellular reprogramming operating at the physiological, cytological and transcriptome level, but also at the chromatin level. A number of observations point to large-scale chromatin reorganization events associated with cellular differentiation of the female spore mother cells and of the female gametes. These include a reorganization of the heterochromatin compartment, the genome-wide alteration of the histone modification landscape, and the remodeling of nucleosome composition. The dynamic expression of DNA methyltransferases and actors of small RNA pathways also suggest additional, global epigenetic alterations that remain to be characterized. Are these events a cause or a consequence of cellular differentiation, and how do they contribute to cell fate transition? Does chromatin dynamics induce competence for immediate cellular functions (meiosis, fertilization), or does it also contribute long-term effects in cellular identity and developmental competence of the reproductive lineage? This review attempts to review these fascinating questions.
Subject(s)
Chromatin/metabolism , Ovule/physiology , Plant Cells/physiology , Cell Differentiation , Chromatin/genetics , Chromatin/ultrastructure , DNA Methylation , Epigenesis, Genetic , Gene Expression Regulation, Plant , MeiosisABSTRACT
The life cycle of flowering plants is marked by several post-embryonic developmental transitions during which novel cell fates are established. Notably, the reproductive lineages are first formed during flower development. The differentiation of spore mother cells, which are destined for meiosis, marks the somatic-to-reproductive fate transition. Meiosis entails the formation of the haploid multicellular gametophytes, from which the gametes are derived, and during which epigenetic reprogramming takes place. Here we show that in the Arabidopsis female megaspore mother cell (MMC), cell fate transition is accompanied by large-scale chromatin reprogramming that is likely to establish an epigenetic and transcriptional status distinct from that of the surrounding somatic niche. Reprogramming is characterized by chromatin decondensation, reduction in heterochromatin, depletion of linker histones, changes in core histone variants and in histone modification landscapes. From the analysis of mutants in which the gametophyte fate is either expressed ectopically or compromised, we infer that chromatin reprogramming in the MMC is likely to contribute to establishing postmeiotic competence to the development of the pluripotent gametophyte. Thus, as in primordial germ cells of animals, the somatic-to-reproductive cell fate transition in plants entails large-scale epigenetic reprogramming.
Subject(s)
Arabidopsis/cytology , Arabidopsis/genetics , Chromatin/genetics , Arabidopsis/physiology , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Histones/genetics , Histones/metabolism , Reproduction/genetics , Reproduction/physiologyABSTRACT
Genomic imprinting results in monoallelic gene expression in a parent-of-origin-dependent manner and is regulated by the differential epigenetic marking of the parental alleles. In plants, genomic imprinting has been primarily described for genes expressed in the endosperm, a tissue nourishing the developing embryo that does not contribute to the next generation. In Arabidopsis, the genes MEDEA (MEA) and PHERES1 (PHE1), which are imprinted in the endosperm, are also expressed in the embryo; whether their embryonic expression is regulated by imprinting or not, however, remains controversial. In contrast, the maternally expressed in embryo 1 (mee1) gene of maize is clearly imprinted in the embryo. We identified several imprinted candidate genes in an allele-specific transcriptome of hybrid Arabidopsis embryos and confirmed parent-of-origin-dependent, monoallelic expression for eleven maternally expressed genes (MEGs) and one paternally expressed gene (PEG) in the embryo, using allele-specific expression analyses and reporter gene assays. Genetic studies indicate that the Polycomb Repressive Complex 2 (PRC2) but not the DNA METHYLTRANSFERASE1 (MET1) is involved in regulating imprinted expression in the embryo. In the seedling, all embryonic MEGs and the PEG are expressed from both parents, suggesting that the imprint is erased during late embryogenesis or early vegetative development. Our finding that several genes are regulated by genomic imprinting in the Arabidopsis embryo clearly demonstrates that this epigenetic phenomenon is not a unique feature of the endosperm in both monocots and dicots.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , Genomic Imprinting , Repressor Proteins/genetics , Seeds/genetics , Alleles , Arabidopsis/growth & development , Arabidopsis Proteins/biosynthesis , DNA (Cytosine-5-)-Methyltransferases/genetics , DNA Methylation/genetics , Endosperm/genetics , Endosperm/growth & development , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , MADS Domain Proteins/biosynthesis , Magnoliopsida/genetics , Polycomb Repressive Complex 2 , Repressor Proteins/biosynthesis , Seeds/growth & development , Zea mays/geneticsABSTRACT
The proper balance of parental genomic contributions to the fertilized embryo and endosperm is essential for their normal growth and development. The characterization of many gametophytic maternal effect (GME) mutants affecting seed development indicates that there are certain classes of genes with a predominant maternal contribution. We present a detailed analysis of the GME mutant zak ixik (zix), which displays delayed and arrested growth at the earliest stages of embryo and endosperm development. ZIX encodes an Armadillo repeat (Arm) protein highly conserved across eukaryotes. Expression studies revealed that ZIX manifests a GME through preferential maternal expression in the early embryo and endosperm. This parent-of-origin-dependent expression is regulated by neither the histone and DNA methylation nor the DNA demethylation pathways known to regulate some other GME mutants. The ZIX protein is localized in the cytoplasm and nucleus of cells in reproductive tissues and actively dividing root zones. The maternal ZIX allele is required for the maternal expression of miniseed3. Collectively, our results reveal a reproductive function of plant Arm proteins in promoting early seed growth, which is achieved through a distinct GME of ZIX that involves mechanisms for maternal allele-specific expression that are independent of the well-established pathways.
Subject(s)
Arabidopsis Proteins/physiology , Arabidopsis/growth & development , Armadillo Domain Proteins/physiology , Seeds/growth & development , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Armadillo Domain Proteins/genetics , Armadillo Domain Proteins/metabolism , Cell Nucleus/metabolism , Cytoplasm/metabolism , Endosperm/genetics , Endosperm/growth & development , Endosperm/metabolism , Gene Expression Regulation, Plant , Plant Roots/metabolism , Seeds/genetics , Seeds/metabolism , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
Genomic imprinting results in monoallelic gene expression in a parent-of-origin-dependent manner. It is achieved by the differential epigenetic marking of parental alleles. Over the past decade, studies in the model systems Arabidopsis thaliana and maize (Zea mays) have shown a strong correlation between silent or active states with epigenetic marks, such as DNA methylation and histone modifications, but the nature of the primary imprint has not been clearly established for all imprinted genes. Phenotypes and expression patterns of imprinted genes have fueled the perception that genomic imprinting is specific to the endosperm, a seed tissue that does not contribute to the next generation. However, several lines of evidence suggest a potential role for imprinting in the embryo, raising questions as to how imprints are erased and reset from one generation to the next. Imprinting regulation in flowering plants shows striking similarities, but also some important differences, compared with the mechanisms of imprinting described in mammals. For example, some imprinted genes are involved in seed growth and viability in plants, which is similar in mammals, where imprinted gene regulation is essential for embryonic development. However, it seems to be more flexible in plants, as imprinting requirements can be bypassed to allow the development of clonal offspring in apomicts.
Subject(s)
Epigenomics , Genomic Imprinting , Seeds/genetics , Animals , Arabidopsis/genetics , DNA Methylation , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Genes, Plant , Histones/genetics , Histones/metabolism , Seeds/growth & development , Zea mays/geneticsABSTRACT
Germ line specification is a crucial step in the life cycle of all organisms. For sexual plant reproduction, the megaspore mother cell (MMC) is of crucial importance: it marks the first cell of the plant "germline" lineage that gets committed to undergo meiosis. One of the meiotic products, the functional megaspore, subsequently gives rise to the haploid, multicellular female gametophyte that harbours the female gametes. The MMC is formed by selection and differentiation of a single somatic, sub-epidermal cell in the ovule. The transcriptional network underlying MMC specification and differentiation is largely unknown. We provide the first transcriptome analysis of an MMC using the model plant Arabidopsis thaliana with a combination of laser-assisted microdissection and microarray hybridizations. Statistical analyses identified an over-representation of translational regulation control pathways and a significant enrichment of DEAD/DEAH-box helicases in the MMC transcriptome, paralleling important features of the animal germline. Analysis of two independent T-DNA insertion lines suggests an important role of an enriched helicase, MNEME (MEM), in MMC differentiation and the restriction of the germline fate to only one cell per ovule primordium. In heterozygous mem mutants, additional enlarged MMC-like cells, which sometimes initiate female gametophyte development, were observed at higher frequencies than in the wild type. This closely resembles the phenotype of mutants affected in the small RNA and DNA-methylation pathways important for epigenetic regulation. Importantly, the mem phenotype shows features of apospory, as female gametophytes initiate from two non-sister cells in these mutants. Moreover, in mem gametophytic nuclei, both higher order chromatin structure and the distribution of LIKE HETEROCHROMATIN PROTEIN1 were affected, indicating epigenetic perturbations. In summary, the MMC transcriptome sets the stage for future functional characterization as illustrated by the identification of MEM, a novel gene involved in the restriction of germline fate.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis , Genome, Plant , Germ Cells, Plant/metabolism , Ovule/genetics , RNA Helicases , Stem Cells/metabolism , Transcriptome/genetics , Arabidopsis/genetics , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Cell Differentiation/genetics , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Epigenesis, Genetic , Gene Expression Profiling , Gene Expression Regulation, Plant , Germ Cells, Plant/cytology , In Situ Hybridization , Meiosis , Microdissection , Mutagenesis, Insertional , Oligonucleotide Array Sequence Analysis , Ovule/metabolism , Phylogeny , Protein Biosynthesis , RNA Helicases/genetics , RNA Helicases/metabolism , Stem Cells/cytologyABSTRACT
Whether deposited maternal products are important during early seed development in flowering plants remains controversial. Here, we show that RNA interference-mediated downregulation of transcription is deleterious to endosperm development but does not block zygotic divisions. Furthermore, we show that RNA POLYMERASE II is less active in the embryo than in the endosperm. This dimorphic pattern is established late during female gametogenesis and is inherited by the two products of fertilization. This juxtaposition of distinct transcriptional activities correlates with differential patterns of histone H3 lysine 9 dimethylation, LIKE HETEROCHROMATIN PROTEIN1 localization, and Histone H2B turnover in the egg cell versus the central cell. Thus, distinct epigenetic and transcriptional patterns in the embryo and endosperm are already established in their gametic progenitors. We further demonstrate that the non-CG DNA methyltransferase CHROMOMETHYLASE3 (CMT3) and DEMETER-LIKE DNA glycosylases are required for the correct distribution of H3K9 dimethylation in the egg and central cells, respectively, and that plants defective for CMT3 activity show abnormal embryo development. Our results provide evidence that cell-specific mechanisms lead to the differentiation of epigenetically distinct female gametes in Arabidopsis thaliana. They also suggest that the establishment of a quiescent state in the zygote may play a role in the reprogramming of the young plant embryo.
Subject(s)
Arabidopsis/genetics , Chromatin/metabolism , Ovule , Seeds/metabolism , Transcription, Genetic , Arabidopsis/embryologyABSTRACT
In mammals and seed plants, a subset of genes is regulated by genomic imprinting where an allele's activity depends on its parental origin. The parental conflict theory suggests that genomic imprinting evolved after the emergence of an embryo-nourishing tissue (placenta and endosperm), resulting in an intragenomic parental conflict over the allocation of nutrients from mother to offspring. It was predicted that imprinted genes, which arose through antagonistic co-evolution driven by a parental conflict, should be subject to positive darwinian selection. Here we show that the imprinted plant gene MEDEA (MEA), which is essential for seed development, originated during a whole-genome duplication 35 to 85 million years ago. After duplication, MEA underwent positive darwinian selection consistent with neo-functionalization and the parental conflict theory. MEA continues to evolve rapidly in the out-crossing species Arabidopsis lyrata but not in the self-fertilizing species Arabidopsis thaliana, where parental conflicts are reduced. The paralogue of MEA, SWINGER (SWN; also called EZA1), is not imprinted and evolved under strong purifying selection because it probably retained the ancestral function of the common precursor gene. The evolution of MEA suggests a late origin of genomic imprinting within the Brassicaceae, whereas imprinting is thought to have originated early within the mammalian lineage.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , Evolution, Molecular , Genes, Plant/genetics , Genomic Imprinting/genetics , Selection, Genetic , Alleles , Animals , Arabidopsis/embryology , Arabidopsis/growth & development , Gene Duplication , Gene Expression Regulation, Plant , Genome, Plant/genetics , Mammals/genetics , Models, Genetic , Molecular Sequence DataABSTRACT
Introduction: Differentiation of spore mother cells marks the somatic-to-reproductive transition in higher plants. Spore mother cells are critical for fitness because they differentiate into gametes, leading to fertilization and seed formation. The female spore mother cell is called the megaspore mother cell (MMC) and is specified in the ovule primordium. The number of MMCs varies by species and genetic background, but in most cases, only a single mature MMC enters meiosis to form the embryo sac. Multiple candidate MMC precursor cells have been identified in both rice and Arabidopsis, so variability in MMC number is likely due to conserved early morphogenetic events. In Arabidopsis, the restriction of a single MMC per ovule, or MMC singleness, is determined by ovule geometry. To look for potential conservation of MMC ontogeny and specification mechanisms, we undertook a morphogenetic description of ovule primordium growth at cellular resolution in the model crop maize. Methods: We generated a collection of 48 three-dimensional (3D) ovule primordium images for five developmental stages, annotated for 11 cell types. Quantitative analysis of ovule and cell morphological descriptors allowed the reconstruction of a plausible developmental trajectory of the MMC and its neighbors. Results: The MMC is specified within a niche of enlarged, homogenous L2 cells, forming a pool of candidate archesporial (MMC progenitor) cells. A prevalent periclinal division of the uppermost central archesporial cell formed the apical MMC and the underlying cell, a presumptive stack cell. The MMC stopped dividing and expanded, acquiring an anisotropic, trapezoidal shape. By contrast, periclinal divisions continued in L2 neighbor cells, resulting in a single central MMC. Discussion: We propose a model where anisotropic ovule growth in maize drives L2 divisions and MMC elongation, coupling ovule geometry with MMC fate.