ABSTRACT
Plant embryogenesis initiates with the establishment of an apical-basal axis; however, the molecular mechanisms accompanying this early event remain unclear. Here, we show that a small cysteine-rich peptide family is required for formation of the zygotic basal cell lineage and proembryo patterning in Arabidopsis. EMBRYO SURROUNDING FACTOR 1 (ESF1) peptides accumulate before fertilization in central cell gametes and thereafter in embryo-surrounding endosperm cells. Biochemical and structural analyses revealed cleavage of ESF1 propeptides to form biologically active mature peptides. Further, these peptides act in a non-cell-autonomous manner and synergistically with the receptor-like kinase SHORT SUSPENSOR to promote suspensor elongation through the YODA mitogen-activated protein kinase pathway. Our findings demonstrate that the second female gamete and its sexually derived endosperm regulate early embryonic patterning in flowering plants.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/embryology , Body Patterning , Flowers/embryology , Seeds/embryology , Amino Acid Sequence , Arabidopsis/genetics , Arabidopsis Proteins/chemistry , Arabidopsis Proteins/genetics , Endosperm/embryology , Endosperm/genetics , Flowers/genetics , Gene Duplication , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Gene Knockout Techniques , Interleukin-1 Receptor-Associated Kinases/metabolism , MAP Kinase Kinase Kinases/metabolism , Molecular Sequence Data , Peptides/chemistry , Peptides/genetics , Peptides/metabolism , Seeds/geneticsABSTRACT
Plant genomes contain several hundred defensin-like (DEFL) genes that encode short cysteine-rich proteins resembling defensins, which are well known antimicrobial polypeptides. Little is known about the expression patterns or functions of many DEFLs because most were discovered recently and hence are not well represented on standard microarrays. We designed a custom Affymetrix chip consisting of probe sets for 317 and 684 DEFLs from Arabidopsis thaliana and Medicago truncatula, respectively for cataloging DEFL expression in a variety of plant organs at different developmental stages and during symbiotic and pathogenic associations. The microarray analysis provided evidence for the transcription of 71% and 90% of the DEFLs identified in Arabidopsis and Medicago, respectively, including many of the recently annotated DEFL genes that previously lacked expression information. Both model plants contain a subset of DEFLs specifically expressed in seeds or fruits. A few DEFLs, including some plant defensins, were significantly up-regulated in Arabidopsis leaves inoculated with Alternaria brassicicola or Pseudomonas syringae pathogens. Among these, some were dependent on jasmonic acid signaling or were associated with specific types of immune responses. There were notable differences in DEFL gene expression patterns between Arabidopsis and Medicago, as the majority of Arabidopsis DEFLs were expressed in inflorescences, while only a few exhibited root-enhanced expression. By contrast, Medicago DEFLs were most prominently expressed in nitrogen-fixing root nodules. Thus, our data document salient differences in DEFL temporal and spatial expression between Arabidopsis and Medicago, suggesting distinct signaling routes and distinct roles for these proteins in the two plant species.
Subject(s)
Arabidopsis/genetics , Defensins/genetics , Gene Expression Regulation, Plant , Medicago truncatula/genetics , Arabidopsis/microbiology , Cluster Analysis , Gene Expression Profiling/methods , Genome, Plant , Host-Pathogen Interactions/genetics , Organ Specificity/genetics , Plant Diseases/genetics , Plant Diseases/microbiology , Plants, Genetically Modified , Reproducibility of Results , Seedlings/genetics , Signal Transduction , Symbiosis/geneticsABSTRACT
Imprinted genes are commonly expressed in mammalian placentas and in plant seed endosperms, where they exhibit preferential uniparental allelic expression. In mammals, imprinted genes directly regulate placental function and nutrient distribution from mother to fetus; however, none of the >60 imprinted genes thus far reported in plants have been demonstrated to play an equivalent role in regulating the flow of resources to the embryo. Here we show that imprinted Maternally expressed gene1 (Meg1) in maize is both necessary and sufficient for the establishment and differentiation of the endosperm nutrient transfer cells located at the mother:seed interface. Consistent with these findings, Meg1 also regulates maternal nutrient uptake, sucrose partitioning, and seed biomass yield. In addition, we generated an imprinted and nonimprinted synthetic Meg1 ((syn)Meg1) dosage series whereby increased dosage and absence of imprinting both resulted in an unequal investment of maternal resources into the endosperm. These findings highlight dosage regulation by genomic imprinting as being critical for maintaining a balanced distribution of maternal nutrients to filial tissues in plants, as in mammals. However, unlike in mammals, Meg1 is a maternally expressed imprinted gene that surprisingly acts to promote rather than restrict nutrient allocation to the offspring.
Subject(s)
Endosperm/metabolism , Genomic Imprinting , Zea mays/metabolism , Cell Differentiation , Endosperm/cytology , Gene Expression Regulation, Plant , Genes, Plant , Zea mays/cytology , Zea mays/geneticsABSTRACT
Cell-cell communication in plants is essential for the correct co-ordination of reproduction, growth, and development. Studies to dissect this mode of communication have previously focussed primarily on the action of plant hormones as mediators of intercellular signalling. In animals, peptide signalling is a well-documented intercellular communication system, however, relatively little is known about this system in plants. In recent years, numerous reports have emerged about small, secreted peptides controlling different aspects of plant reproduction. Interestingly, most of these peptides are cysteine-rich, and there is convincing evidence suggesting multiple roles for related cysteine-rich peptides (CRPs) as signalling factors in developmental patterning as well as during plant pathogen responses and symbiosis. In this review, we discuss how CRPs are emerging as key signalling factors in regulating multiple aspects of vegetative growth and reproductive development in plants.
Subject(s)
Cell Communication/physiology , Cysteine/metabolism , Gene Expression Regulation, Plant , Peptides/metabolism , Plant Development , Bacteria/growth & development , Peptides/chemistry , Peptides/genetics , Plant Growth Regulators/physiology , Plant Immunity/physiology , Plant Proteins/chemistry , Plant Proteins/genetics , Plant Proteins/metabolism , Plants/metabolism , Plants/microbiology , Reproduction/physiology , Signal Transduction/physiology , Symbiosis/physiologyABSTRACT
BACKGROUND AND AIMS: The results of wide- or interploidy crosses in angiosperms are unpredictable and often lead to seed abortion. The consequences of reciprocal interploidy crosses have been explored in maize in detail, focusing on alterations to tissue domains in the maize endosperm, and changes in endosperm-specific gene expression. METHODS: Following reciprocal interploidy crosses between diploid and tetraploid maize lines, development of endosperm domains was studied using GUS reporter lines, and gene expression in resulting kernels was investigated using semi-quantitative RT-PCR on endosperms isolated at different stages of development. KEY RESULTS: Reciprocal interploidy crosses result in very small, largely infertile seeds with defective endosperms. Seeds with maternal genomic excess are smaller than those with paternal genomic excess, their endosperms cellularize earlier and they accumulate significant quantities of starch. Endosperms from the reciprocal cross undergo an extended period of cell proliferation, and accumulate little starch. Analysis of reporter lines and gene expression studies confirm that functional domains of the endosperm are severely disrupted, and are modified differently according to the direction of the interploidy cross. CONCLUSIONS: Interploidy crosses affect factors which regulate the balance between cell proliferation and cell differentiation within the endosperm. In particular, unbalanced crosses in maize affect transfer cell differentiation, and lead to the temporal deregulation of the ontogenic programme of endosperm development.
Subject(s)
Crosses, Genetic , Genome, Plant , Ploidies , Seeds/growth & development , Zea mays/embryology , Genes, Reporter , Reverse Transcriptase Polymerase Chain Reaction , Starch/metabolism , Zea mays/geneticsABSTRACT
Viviparous1 (Vp1) encodes a B3 domain-containing transcription factor that is a key regulator of seed maturation in maize (Zea mays). However, the mechanisms of Vp1 regulation are not well understood. To examine physiological factors that may regulate Vp1 expression, transcript levels were monitored in maturing embryos placed in culture under different conditions. Expression of Vp1 decreased after culture in hormone-free medium, but was induced by salinity or osmotic stress. Application of exogenous abscisic acid (ABA) also induced transcript levels within 1 h in a dose-dependent manner. The Vp1 promoter fused to beta-glucuronidase or green fluorescent protein reproduced the endogenous Vp1 expression patterns in transgenic maize plants and also revealed previously unknown expression domains of Vp1. The Vp1 promoter is active in the embryo and aleurone cells of developing seeds and, upon drought stress, was also found in phloem cells of vegetative tissues, including cobs, leaves, and stems. Sequence analysis of the Vp1 promoter identified a potential ABA-responsive complex, consisting of an ACGT-containing ABA response element (ABRE) and a coupling element 1-like motif. Electrophoretic mobility shift assay confirmed that the ABRE and putative coupling element 1 components specifically bound proteins in embryo nuclear protein extracts. Treatment of embryos in hormone-free Murashige and Skoog medium blocked the ABRE-protein interaction, whereas exogenous ABA or mannitol treatment restored this interaction. Our data support a model for a VP1-dependent positive feedback mechanism regulating Vp1 expression during seed maturation.
Subject(s)
Abscisic Acid/pharmacology , Gene Expression Regulation, Plant/drug effects , Plant Proteins/genetics , Seeds/drug effects , Signal Transduction/drug effects , Zea mays/genetics , Zea mays/metabolism , Gene Expression Regulation, Developmental/drug effects , Genes, Reporter , Glucuronidase/metabolism , Plant Growth Regulators/pharmacology , Plant Proteins/metabolism , Plants, Genetically Modified , Promoter Regions, Genetic , Seeds/genetics , Seeds/metabolism , Zea mays/cytologyABSTRACT
The pentatricopeptide repeat (PPR) family represents one of the largest gene families in plants, with >440 members annotated in Arabidopsis thaliana. PPR proteins are thought to have a major role in the regulation of posttranscriptional processes in organelles. Recent studies have shown that Arabidopsis PPR proteins play an essential, nonredundant role during embryogenesis. Here, we demonstrate that mutations in empty pericarp4 (emp4), a maize (Zea mays) PPR-encoding gene, confer a seed-lethal phenotype. Mutant endosperms are severely impaired, with highly irregular differentiation of transfer cells in the nutrient-importing basal endosperm. Analysis of homozygous mutant plants generated from embryo-rescue experiments indicated that emp4 also affects general plant growth. The emp4-1 mutation was identified in an active Mutator (Mu) population, and cosegregation analysis revealed that it arose from a Mu3 element insertion. Evidence of emp4 molecular cloning was provided by the isolation of four additional emp4 alleles obtained by a reverse genetics approach. emp4 encodes a novel type of PPR protein of 614 amino acids. EMP4 contains nine 35-amino acid PPR motifs and an N-terminal mitochondrion-targeted sequence peptide, which was confirmed by a translational EMP4-green fluorescent protein fusion that localized to mitochondria. Molecular analyses further suggest that EMP4 is necessary to regulate the correct expression of a small subset of mitochondrial transcripts in the endosperm.
Subject(s)
Plant Proteins/physiology , Zea mays/growth & development , Amino Acid Motifs , Cloning, Molecular , Green Fluorescent Proteins/analysis , Mitochondria/metabolism , Mitochondrial Proteins/genetics , Mitochondrial Proteins/metabolism , Molecular Sequence Data , Multigene Family/physiology , Mutation , Phenotype , Plant Proteins/chemistry , Plant Proteins/genetics , RNA, Messenger/metabolism , Recombinant Fusion Proteins/analysis , Seeds/anatomy & histology , Seeds/growth & development , Seeds/metabolism , Sequence Alignment , Zea mays/genetics , Zea mays/metabolismABSTRACT
In angiosperms, double fertilization of an egg cell and a central cell with two sperm cells results in the formation of a seed containing a diploid embryo and a triploid endosperm. The extent to which the embryo sac controls postfertilization events in the seed is unknown. The novel gametophytic maternal-effect maize mutation, baseless1 (bsl1) affects central cell development within the embryo sac, frequently by altering the position of the two polar nuclei. Despite this irregularity, fertilization is as efficient as in wild type. The spatial expression of basal endosperm-specific transcripts is altered in free-nuclear and cellular mutant endosperms. At later stages of seed development, bsl1 predominantly affects development of the basal endosperm transfer layer (BETL). When bsl1/+ diploid plants were pollinated by wild-type tetraploid plants, the BETL abnormalities observed in bsl1/bsl1/+/+ tetraploid endosperms were diverse and of variable severity. Moreover, the frequency of kernels with severely perturbed BETL development correlated with the percentage of severely affected bsl1 central cells. Therefore, BSL1 is likely required in the central cell before fertilization for correct BETL patterning to occur. These findings provide new genetic evidence that a maternal gametophytic component is necessary for correct endosperm patterning.
Subject(s)
Genes, Plant/physiology , Phenotype , Zea mays/embryology , Zea mays/growth & development , Zea mays/genetics , Gametogenesis/genetics , Gene Expression Regulation, Plant , Models, Biological , Mutation/physiology , Plant Proteins/genetics , Plant Proteins/metabolism , Plants, Genetically Modified , Pollen/embryology , Pollen/genetics , Seeds/embryology , Seeds/genetics , Tissue DistributionABSTRACT
Plant imprinted genes show parent-of-origin expression in seed endosperm, but little is known about the nature of parental imprints in gametes before fertilization. We show here that single differentially methylated regions (DMRs) correlate with allele-specific expression of two maternally expressed genes in the seed and that one DMR is differentially methylated between gametes. Thus, plants seem to have developed similar strategies as mammals to epigenetically mark imprinted genes.
Subject(s)
Epigenesis, Genetic , Genomic Imprinting , Plants/genetics , CpG Islands , DNA Methylation , DNA, Plant/chemistry , DNA, Plant/genetics , Gene Expression Regulation, Plant , Germ Cells/metabolism , Plants/embryology , Plants, Genetically Modified , Zea mays/geneticsABSTRACT
The analysis of cell type-specific gene expression is an essential step in understanding certain biological processes during plant development, such as differentiation. Although methods for isolating specific cell types have been established, the application of cDNA subtraction to small populations of isolated cell types for direct identification of specific or differentially expressed transcripts has not yet been reported. As a first step in the identification of genes expressed differentially between maize egg cells and central cells, we have manually isolated these types of cell, and applied a suppression-subtractive hybridization (SSH) strategy. After microarray screening of 1030 cDNAs obtained from the subtracted libraries, we identified 340 differentially expressed clones. Of these, 142 were sequenced, which resulted in the identification of 62 individual cDNAs. The expression patterns of 20 cDNAs were validated by quantitative RT-PCR, through which we identified five transcripts with cell type-specific expression. The specific localization of some of these transcripts was also confirmed by in situ hybridization on embryo sac sections. Taken together, our data demonstrate the effectiveness of our approach in identifying differentially expressed and cell type-specific transcripts of relatively low abundance. This was also confirmed by the identification of previously reported egg cell- and central cell-specific genes in our screen. Importantly, from our analysis we identified a significant number of novel sequences not present in other embryo sac or, indeed, in other plant expressed sequence tag (EST) databases. Thus, in combination with standard EST sequencing and microarray hybridization strategies, our approach of differentially screening subtracted cDNAs will add substantially to the expression information in spatially highly resolved transcriptome analyses.
Subject(s)
Gene Expression Regulation, Plant/genetics , Gene Library , Ovum/metabolism , Zea mays/cytology , Zea mays/genetics , Gene Expression Profiling , Gene Expression Regulation, Developmental/genetics , Molecular Sequence Data , Seeds/metabolism , Zea mays/growth & developmentABSTRACT
A novel stigma-specific class III peroxidase gene, SSP (Stigma-Specific Peroxidase), has been isolated from the self-incompatible daisy Senecio squalidus L. (Asteraceae). Expression of SSP in flower buds is developmentally regulated, with maximal levels of expression coinciding with anthesis, when stigmas are most receptive to pollen and when self-incompatibility is fully developed. In situ hybridization revealed SSP expression to be localized exclusively to the specialized secretory epidermal cells (papillae) of the stigma, which receive and discriminate pollen. SSP is therefore the first tissue-specific and cell-specific peroxidase gene identified in a plant. SSP belongs to a distinct clade of class III plant peroxidases that possess two introns, instead of the more normal situation of three conserved introns. The deduced amino acid sequence of SSP revealed a 27 amino acid signal peptide, suggesting that the SSP protein is secreted to the cell wall of the stigmatic papillae. In-gel peroxidase activity assays showed that SSP has relatively low peroxidase activity compared to other, as yet uncharacterized, peroxidases present in stigmatic extracts. Six SSP alleles have been cloned from different lines of S. squalidus carrying a range of self-incompatibility (S)-alleles but there was no consistent association between the presence of a particular SSP allele and S-genotype indicating that SSP is not the female determinant of SSI in S. squalidus. Nevertheless, the precise expression of SSP in stigmatic papillae suggests that it may have a more general function in pollen-stigma interactions, or alternatively in protection of stigmas from pathogen attack. Extensive database screens have identified homologues of SSP in other plant species, but available expression data for these genes indicates that none are flower-specific, suggesting that SSP represents a new functional type of class III peroxidase specific to the stigma. We discuss the possible function(s) of S. squalidus SSP in pollen-stigma interactions and in protection of stigmas from pathogen attack.
Subject(s)
Flowers/enzymology , Peroxidase/genetics , Senecio/genetics , Alleles , Amino Acid Sequence , Base Sequence , Blotting, Northern , Blotting, Southern , Cloning, Molecular , DNA, Complementary/chemistry , DNA, Complementary/genetics , DNA, Complementary/isolation & purification , DNA, Plant/chemistry , DNA, Plant/isolation & purification , Fertility/genetics , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Plant , Genotype , In Situ Hybridization , Isoelectric Focusing , Isoenzymes/chemistry , Isoenzymes/genetics , Isoenzymes/metabolism , Molecular Sequence Data , Peroxidase/chemistry , Peroxidase/metabolism , Phylogeny , Polymorphism, Genetic , RNA, Plant/genetics , RNA, Plant/metabolism , Senecio/enzymology , Sequence Analysis, DNA , Sequence Analysis, Protein , Sequence Homology, Amino AcidABSTRACT
Growth of the maize (Zea mays) endosperm is tightly regulated by maternal zygotic and sporophytic genes, some of which are subject to a parent-of-origin effect. We report here a novel gene, maternally expressed gene1 (meg1), which shows a maternal parent-of-origin expression pattern during early stages of endosperm development but biallelic expression at later stages. Interestingly, a stable reporter fusion containing the meg1 promoter exhibits a similar pattern of expression. meg1 is exclusively expressed in the basal transfer region of the endosperm. Further, we show that the putatively processed MEG1 protein is glycosylated and subsequently localized to the labyrinthine ingrowths of the transfer cell walls. Hence, the discovery of a parent-of-origin gene expressed solely in the basal transfer region opens the door to epigenetic mechanisms operating in the endosperm to regulate certain aspects of nutrient trafficking from the maternal tissue into the developing seed.
Subject(s)
Gene Expression Regulation, Plant , Genes, Plant/genetics , Glycoproteins/genetics , Zea mays/genetics , Amino Acid Sequence , Cloning, Molecular , Glucuronidase/genetics , Models, Molecular , Molecular Sequence Data , Multigene Family , Plant Proteins/genetics , Plants, Genetically Modified , Protein Conformation , Reproduction , Sequence Alignment , Sequence Homology, Amino Acid , Zygote/physiologyABSTRACT
Cereal endosperm tissues account for most of the world's calorific intake, yet the regulation of monocot seed development remains poorly understood. The maize endosperm originates with a series of free-nuclear divisions, followed by cellularisation and subsequent formation of a range of functional cellular domains. We describe the isolation and characterisation of a mutation that induces aberrant globular embryo and endosperm morphology, globby1-1 (glo1-1). Our data indicate that glo1-1 plays a role in nuclear division and cytokinesis in the developing seed. Pattern formation in the embryo is severely impaired with development arresting at premature stages, while in the endosperm, the effects of the glo1-1 mutation are manifest at the free-nuclear or syncytial stage. During cellularisation, and at later stages of development, aberrant cell division and localised domains of cell proliferation are apparent in glo1-1 endosperms. As a consequence, cell fate acquisition and subsequent differentiation of endosperm tissues are affected to varying degrees of severity. To date, it has been hypothesised that BETL cell fate is specified in the syncytium and that cell files subsequently develop in response to a gradient of signal(s) derived from the maternal pedicel region. Based on our findings, however, we propose that specification of BETL cells is an irreversible event that occurs within a narrow window of syncytial development, and that BETL cell identity is subsequently inherited in a lineage-dependent manner. Additionally, our data suggest that acquisition of aleurone cell fate does not solely rely upon signalling from the maternal surrounding tissue to the periphery of the endosperm, as previously thought, but that other factor(s) present within the endosperm are involved.