RESUMO
Animal fertilization relies on hundreds of sperm racing toward the egg, whereas, in angiosperms, only two sperm cells are delivered by a pollen tube to the female gametes (egg cell and central cell) for double fertilization. However, unsuccessful fertilization under this one-pollen-tube design can be detrimental to seed production and plant survival. To mitigate this risk, unfertilized-gamete-controlled extra pollen tube entry has been evolved to bring more sperm cells and salvage fertilization. Despite its importance, the underlying molecular mechanism of this phenomenon remains unclear. In this study, we report that, in Arabidopsis, the central cell secretes peptides SALVAGER1 and SALVAGER2 in a directional manner to attract pollen tubes when the synergid-dependent attraction fails or is terminated by pollen tubes carrying infertile sperm cells. Moreover, loss of SALs impairs the fertilization recovery capacity of the ovules. Therefore, this research uncovers a female gamete-attraction system that salvages seed production for reproductive assurance.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Animais , Arabidopsis/fisiologia , Fertilização , Tubo Polínico , Sementes , Células Germinativas VegetaisRESUMO
Flowers have a species-specific fertile period during which pollination and fertilization have to occur to initiate seed and fruit development. Unpollinated flowers remain receptive for mere hours in some species, and up to several weeks in others before flower senescence terminates fertility. As such, floral longevity is a key trait subject to both natural selection and plant breeding. Within the flower, the life span of the ovule containing the female gametophyte is decisive for fertilization and the initiation of seed development. Here, we show that unfertilized ovules in Arabidopsis thaliana undergo a senescence program that generates morphological and molecular hallmarks of canonical programmed cell death processes in the sporophytically derived ovule integuments. Transcriptome profiling of isolated aging ovules revealed substantial transcriptomic reprogramming during ovule senescence, and identified up-regulated transcription factors as candidate regulators of these processes. Combined mutation of three most-up-regulated NAC (NAM, ATAF1/2, and CUC2) transcription factors, NAP/ANAC029, SHYG/ANAC047, and ORE1/ANAC092, caused a substantial delay in ovule senescence and an extension of fertility in Arabidopsis ovules. These results suggest that timing of ovule senescence and duration of gametophyte receptivity are subject to genetic regulation controlled by the maternal sporophyte.
Assuntos
Arabidopsis , Fatores de Transcrição , Óvulo Vegetal , Melhoramento Vegetal , FertilidadeRESUMO
Arabidopsis mitochondria-targeted heat shock protein 70 (mtHSC70-1) plays important roles in the establishment of cytochrome c oxidase-dependent respiration and redox homeostasis during the vegetative growth of plants. Here, we report that knocking out the mtHSC70-1 gene led to a decrease in plant fertility; the fertility defect of the mutant was completely rescued by introducing the mtHSC70-1 gene. mtHSC70-1 mutants also showed defects in female gametophyte (FG) development, including delayed mitosis, abnormal nuclear position, and ectopic gene expression in the embryo sacs. In addition, we found that an Arabidopsis mitochondrial J-protein gene (DjA30) mutant, j30+/- , had defects in FG development and fertility similar to those of mtHSC70-1 mutant. mtHSC70-1 and DjA30 had similar expression patterns in FGs and interacted in vivo, suggesting that these two proteins might cooperate during female gametogenesis. Further, respiratory chain complex IV activity in mtHSC70-1 and DjA30 mutant embryo sacs was markedly downregulated; this led to the accumulation of mitochondrial reactive oxygen species (ROS). Scavenging excess ROS by introducing Mn-superoxide dismutase 1 or catalase 1 gene into the mtHSC70-1 mutant rescued FG development and fertility. Altogether, our results suggest that mtHSC70-1 and DjA30 are essential for the maintenance of ROS homeostasis in the embryo sacs and provide direct evidence for the roles of ROS homeostasis in embryo sac maturation and nuclear patterning, which might determine the fate of gametic and accessory cells.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Óvulo Vegetal/genética , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Regulação da Expressão Gênica de PlantasRESUMO
In gymnosperms such as Ginkgo biloba, the arrival of pollen plays a key role in ovule development, before fertilization occurs. Accordingly, G. biloba female plants geographically isolated from male plants abort all their ovules after the pollination drop emission, which is the event that allows the ovule to capture pollen grains. To decipher the mechanism induced by pollination required to avoid ovule senescence and then abortion, we compared the transcriptomes of pollinated and unpollinated ovules at three time points after the end of the emission of pollination drop. Transcriptomic and in situ expression analyses revealed that several key genes involved in programmed cell death such as senescence and apoptosis, DNA replication, and cell cycle regulation were differentially expressed in unpollinated ovules compared to pollinated ovules. We provide evidence that the pollen captured by the pollination drop affects auxin local accumulation and might cause deregulation of key genes required for the ovule's programmed cell death, activating both the cell cycle regulation and DNA replication genes.
Assuntos
Ginkgo biloba , Óvulo Vegetal , Pólen , Polinização , Óvulo Vegetal/crescimento & desenvolvimento , Óvulo Vegetal/fisiologia , Óvulo Vegetal/genética , Pólen/genética , Pólen/crescimento & desenvolvimento , Pólen/fisiologia , Ginkgo biloba/genética , Ginkgo biloba/fisiologia , Ginkgo biloba/crescimento & desenvolvimento , Transcriptoma , Regulação da Expressão Gênica de PlantasRESUMO
A functional female gametophyte is the basis of successful sexual reproduction in flowering plants. During female gametophyte development, the megaspore mother cell (MMC), which differentiates from a single subepidermal somatic cell in the nucellus, undergoes meiosis to produce four megaspores; only the one at the chalazal end, referred to as the functional megaspore (FM), then undergoes three rounds of mitosis and develops into a mature embryo sac. Here, we report that RING1A and RING1B (RING1A/B), two functionally redundant Polycomb proteins in Arabidopsis, are critical for female gametophyte development. Mutations of RING1A/B resulted in defects in the specification of the MMC and the FM, and in the subsequent mitosis of the FM, thereby leading to aborted ovules. Detailed analysis revealed that several genes essential for female gametophyte development were ectopically expressed in the ring1a ring1b mutant, including Argonaute (AGO) family genes and critical transcription factors. Furthermore, RING1A/B bound to some of these genes to promote H2A monoubiquitination (H2Aub). Taken together, our study shows that RING1A/B promote H2Aub modification at key genes for female gametophyte development, suppressing their expression to ensure that the development progresses correctly.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Óvulo Vegetal , Ubiquitinação , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Óvulo Vegetal/crescimento & desenvolvimento , Óvulo Vegetal/genética , Óvulo Vegetal/metabolismo , Complexo Repressor Polycomb 1/metabolismo , Complexo Repressor Polycomb 1/genética , Regulação da Expressão Gênica de Plantas , Histonas/metabolismo , Histonas/genética , Proteínas de TransporteRESUMO
The development of the embryo sac is an important factor that affects seed setting in rice. Numerous genes associated with embryo sac (ES) development have been identified in plants; however, the function of the DEAD-box RNA helicase family genes is poorly known in rice. Here, we characterized a rice DEAD-box protein, RH52A, which is localized in the nucleus and cytoplasm and highly expressed in the floral organs. The knockout mutant rh52a displayed partial ES sterility, including degeneration of the ES (21%) and the presence of a double-female-gametophyte (DFG) structure (11.8%). The DFG developed from two functional megaspores near the chalazal end in one ovule, and 3.4% of DFGs were able to fertilize via the sac near the micropylar pole in rh52a. RH52A was found to interact with MFS1 and ZIP4, both of which play a role in homologous recombination in rice meiosis. RNA-sequencing identified 234 down-regulated differentially expressed genes associated with reproductive development, including two, MSP1 and HSA1b, required for female germline cell specification. Taken together, our study demonstrates that RH52A is essential for the development of the rice embryo sac and provides cytological details regarding the formation of DFGs.
Assuntos
RNA Helicases DEAD-box , Oryza , Proteínas de Plantas , Oryza/genética , Oryza/metabolismo , Oryza/crescimento & desenvolvimento , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genética , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Regulação da Expressão Gênica de Plantas , Óvulo Vegetal/crescimento & desenvolvimento , Óvulo Vegetal/genética , Óvulo Vegetal/metabolismoRESUMO
In angiosperm, two immotile sperm cells are delivered to the female gametes for fertilization by a pollen tube, which perceives guidance cues from ovules at least at two critical sites, micropyle for short-distance guidance and funiculus for comparably longer distance guidance. Compared with the great progress in understanding pollen tube micropylar guidance, little is known about the signaling for funicular guidance. Here, we show that funiculus plays an important role in pollen tube guidance and report that female gametophyte (FG) plays a critical role in funicular guidance by analysis of a 3-dehydroquinate synthase (DHQS) mutant. Loss function of DHQS in FG interrupts pollen tube funicular guidance, suggesting that the guiding signal is generated from FG. We show the evidence that the capacity of funicular guidance is established during FG functional specification after the establishment of cell identity. Specific expression of DHQS in the synergid cells, central cells, or egg cells can rescue funicular guidance defect in dhqs/+, indicating all the female germ unit cells are involved in the funicular guidance. The finding reveals that the attracting signal of pollen tube funicular guidance was generated at a site and stage manner and provides novel clue to locate and search for the signal.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Tubo Polínico , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Óvulo Vegetal/metabolismo , Tubo Polínico/metabolismo , Polinização/fisiologia , Sementes/metabolismoRESUMO
Endosperm is a key nutritive tissue that supports the developing embryo or seedling, and serves as a major nutritional source for human and livestock feed. In sexually-reproducing flowering plants, it generally develops after fertilization. However, autonomous endosperm (AE) formation (i.e. independent of fertilization) is also possible. Recent findings of AE loci/ genes and aberrant imprinting in native apomicts, together with a successful initiation of parthenogenesis in rice and lettuce, have enhanced our understanding of the mechanisms bridging sexual and apomictic seed formation. However, the mechanisms driving AE development are not well understood. This review presents novel aspects related to AE development in sexual and asexual plants underlying stress conditions as the primary trigger for AE. Both application of hormones to unfertilized ovules and mutations that impair epigenetic regulation lead to AE development in sexual Arabidopsis thaliana, which may point to a common pathway for both phenomena. Apomictic-like AE development under experimental conditions can take place due to auxin-dependent gene expression and/or DNA methylation.
Assuntos
Arabidopsis , Asteraceae , Humanos , Endosperma/genética , Epigênese Genética , Sementes , Reprodução , Arabidopsis/genéticaRESUMO
The nucellus tissue in flowering plants provides nutrition for the development of the female gametophyte (FG) and young embryo. The nucellus degenerates as the FG develops, but the mechanism controlling the coupled process of nucellar degeneration and FG expansion remains largely unknown. The degeneration process of the nucellus and spatiotemporal auxin distribution in the developing ovule before fertilization were investigated in Arabidopsis thaliana. Nucellar degeneration before fertilization occurs through vacuolar cell death and in an ordered degeneration fashion. This sequential nucellar degeneration is controlled by the signalling molecule auxin. Auxin efflux plays the core role in precisely controlling the spatiotemporal pattern of auxin distribution in the nucellus surrounding the FG. The auxin efflux carrier PIN1 transports maternal auxin into the nucellus while PIN3/PIN4/PIN7 further delivers auxin to degenerating nucellar cells and concurrently controls FG central vacuole expansion. Notably, auxin concentration and auxin efflux are controlled by the maternal tissues, acting as a key communication from maternal to filial tissue.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Ácidos Indolacéticos , Óvulo Vegetal/metabolismoRESUMO
Eukaryotic cells rely on the accuracy and efficiency of vesicular traffic. In plants, disturbances in vesicular trafficking are well studied in quickly dividing root meristem cells or polar growing root hairs and pollen tubes. The development of the female gametophyte, a unique haploid reproductive structure located in the ovule, has received far less attention in studies of vesicular transport. Key molecules providing the specificity of vesicle formation and its subsequent recognition and fusion with the acceptor membrane are Rab proteins. Rabs are anchored to membranes by covalently linked geranylgeranyl group(s) that are added by the Rab geranylgeranyl transferase (RGT) enzyme. Here we show that Arabidopsis plants carrying mutations in the gene encoding the ß-subunit of RGT (rgtb1) exhibit severely disrupted female gametogenesis and this effect is of sporophytic origin. Mutations in rgtb1 lead to internalization of the PIN1 and PIN3 proteins from the basal membranes to vesicles in provascular cells of the funiculus. Decreased transport of auxin out of the ovule is accompanied by auxin accumulation in tissue surrounding the growing gametophyte. In addition, female gametophyte development arrests at the uni- or binuclear stage in a significant portion of the rgtb1 ovules. These observations suggest that communication between the sporophyte and the developing female gametophyte relies on Rab-dependent vesicular traffic of the PIN1 and PIN3 transporters and auxin efflux out of the ovule.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Ácidos Indolacéticos , Óvulo Vegetal/genética , Tubo PolínicoRESUMO
The developmental process of inflorescence and gametophytes is vital for sexual reproduction in rice. Multiple genes and conserved miRNAs have been characterized to regulate the process. The changes of miRNAs expression during the early development of rice inflorescence remain unknown. In this study, the analysis of miRNAs profiles in the early stage of rice inflorescence development identified 671 miRNAs, including 67 known and 44 novel differentially expressed miRNAs (DEMs). Six distinct clusters of miRNAs expression patterns were detected, and Cluster 5 comprised 110 DEMs, including unconserved, rice-specific osa-miR5506. Overexpression of osa-miR5506 caused pleiotropic abnormalities, including over- or under-developed palea, various numbers of floral organs and spikelet indeterminacy. In addition, the defects of ovaries development were frequently characterized by multiple megasporocytes, ovule-free ovary, megasporocyte degenerated and embryo sac degenerated in the transgenic lines. osa-miR5506 targeted REM transcription factor LOC_Os03g11370. Summarily, these results demonstrated that rice-specific osa-miR5506 plays an essential role in the regulation of floral organ number, spikelet determinacy and female gametophyte development in rice.
Assuntos
Inflorescência/crescimento & desenvolvimento , MicroRNAs/metabolismo , Oryza/metabolismo , Óvulo Vegetal/crescimento & desenvolvimento , Perfilação da Expressão Gênica , Meiose , Oryza/genética , Oryza/crescimento & desenvolvimento , Plantas Geneticamente ModificadasRESUMO
BACKGROUND: Glycosylphosphatidylinositol (GPI) addition is one of the several post-translational modifications to proteins that increase their affinity for membranes. In eukaryotes, the GPI transamidase complex (GPI-T) catalyzes the attachment of pre-assembled GPI anchors to GPI-anchored proteins (GAPs) through a transamidation reaction. A mutation in AtGPI8 (gpi8-2), the putative catalytic subunit of GPI-T in Arabidopsis, is transmitted normally through the female gametophyte (FG), indicating the FG tolerates loss of GPI transamidation. In contrast, gpi8-2 almost completely abolishes male gametophyte (MG) function. Still, the unexpected finding that gpi8-2 FGs function normally requires further investigation. Additionally, specific developmental defects in the MG caused by loss of GPI transamidation remain poorly characterized. RESULTS: Here we investigated the effect of loss of AtPIG-S, another GPI-T subunit, in both gametophytes. Like gpi8-2, we showed that a mutation in AtPIG-S (pigs-1) disrupted synergid localization of LORELEI (LRE), a putative GAP critical for pollen tube reception by the FG. Still, pigs-1 is transmitted normally through the FG. Conversely, pigs-1 severely impaired male gametophyte (MG) function during pollen tube emergence and growth in the pistil. A pPIGS:GFP-PIGS transgene complemented these MG defects and enabled generation of pigs-1/pigs-1 seedlings. However, the pPIGS:GFP-PIGS transgene seemingly failed to rescue the function of AtPIG-S in the sporophyte, as pigs-1/pigs-1, pPIGS:GFP-PIGS seedlings died soon after germination. CONCLUSIONS: Characterization of pigs-1 provided further evidence that the FG tolerates loss of GPI transamidation more than the MG and that the MG compared to the FG may be a better haploid system to study the role of GPI-anchoring. Pigs-1 pollen develops normally and thus represent a tool in which GPI anchor biosynthesis and transamidation of GAPs have been uncoupled, offering a potential way to study free GPI in plant development. While previously reported male fertility defects of GPI biosynthesis mutants could have been due either to loss of GPI or GAPs lacking the GPI anchor, our results clarified that the loss of mature GAPs underlie male fertility defects of GPI-deficient pollen grains, as pigs-1 is defective only in the downstream transamidation step.
Assuntos
Aciltransferases/fisiologia , Arabidopsis/enzimologia , Arabidopsis/crescimento & desenvolvimento , Tubo Polínico/crescimento & desenvolvimento , Aciltransferases/genética , Proteínas de Arabidopsis/metabolismo , Clonagem Molecular , Técnicas de Genotipagem , Glicoproteínas de Membrana/metabolismo , Mutação , Pólen/genética , Subunidades Proteicas/genética , Subunidades Proteicas/fisiologia , Reação em Cadeia da Polimerase em Tempo Real , Nicotiana/genéticaRESUMO
The maturation of male and female gametophytes together with its impact on plant sexual reproduction has not received much attention, and the molecular mechanisms underlying the process are largely unknown. Here, we show that Arabidopsis DEAD-box RNA helicase 29 (RH29) is critical for the functional maturation of both male and female gametophytes. Homozygous rh29 mutants could not be obtained, and heterozygous mutant plants were semi-sterile. Progression of the cell cycle in rh29 female gametophytes was delayed. Delayed pollination experiments showed that rh29 female gametophytes underwent cell-fate specification but were unable to develop into functional gametophytes. Functional specification but not morphogenesis was also disrupted in rh29 male gametophytes, causing defective pollen tube growth in the pistil. RH29 was highly and specifically expressed in gametophytic cells. RH29 shares high amino acid sequence identity with yeast Dbp10p, which partially rescues the aborted-ovules phenotype of rh29/RH29 plants. RH29 is essential for the synthesis of REGULATORY PARTICLE TRIPLE A ATPase 5a (RPT5a), a subunit of the regulatory particle of the 26S proteasome. Our results suggest that gametophyte functional maturation is a necessary process for successful fertilization and that RH29 is essential for the functional maturation of both male and female gametophytes.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Proteínas de Arabidopsis/genética , RNA Helicases DEAD-box/genética , Células Germinativas Vegetais , Mutação , Óvulo Vegetal/genéticaRESUMO
The DnaJ-like zinc finger domain-containing proteins are involved in different aspects of plastid function and development. Some of these proteins were recently reported to have dual subcellular localization in the nucleus and plastids. One member of this family, PSA2 (AT2G34860), was found to localize to the thylakoid lumen and regulate the assembly of photosystem I (PSI). However, PSA2 was also annotated as Embryo sac Development Arrest 3 (EDA3) from the observation that its embryo sac development was arrested at the two-nuclear stage. In this study, we characterized the eda3 mutant, and demonstrated that, as compared with the wild-type (WT) plants, the mutant has shorter siliques, fewer siliques per plant, and fewer seeds per silique. Both aborted and undeveloped ovules were observed in siliques of the mutant. By immunoblot analysis, we found that, different from the chloroplast localization in mature leaves, EDA3 localizes in the nucleus in seeds. A nuclear localization signal was identified from the deduced amino acid sequence of EDA3, and also proved to be sufficient for directing its fusion peptide into the nucleus.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Núcleo Celular/metabolismo , Mutação , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Núcleo Celular/genética , Cloroplastos/genética , Cloroplastos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Sinais de Localização Nuclear , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Transporte Proteico , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Dedos de ZincoRESUMO
In flowering plants, cell-cell communication plays a key role in reproductive success, as both pollination and fertilization require pathways that regulate interactions between many different cell types. Some of the most critical of these interactions are those between the pollen tube (PT) and the embryo sac, which ensure the delivery of sperm cells required for double fertilization. Synergid cells function to attract the PT through secretion of small peptides and in PT reception via membrane-bound proteins associated with the endomembrane system and the cell surface. While many synergid-expressed components regulating PT attraction and reception have been identified, few tools exist to study the localization of membrane-bound proteins and the components of the endomembrane system in this cell type. In this study, we describe the localization and distribution of seven fluorescent markers that labelled components of the secretory pathway in synergid cells of Arabidopsis thaliana. These markers were used in co-localization experiments to investigate the subcellular distribution of the two PT reception components LORELEI, a GPI-anchored surface protein, and NORTIA, a MILDEW RESISTANCE LOCUS O protein, both found within the endomembrane system of the synergid cell. These secretory markers are useful tools for both reproductive and cell biologists, enabling the analysis of membrane-associated trafficking within a haploid cell actively involved in polar transport.
Assuntos
Arabidopsis/metabolismo , Via Secretória , Arabidopsis/citologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Biomarcadores , Membrana Celular/metabolismo , Haploidia , Plantas Geneticamente Modificadas , Tubo Polínico/citologia , Tubo Polínico/metabolismo , Via Secretória/genéticaRESUMO
BACKGROUND: As important female reproductive tissues, the rice (Oryza sativa L.) ovule and female gametophyte is significant in terms of their fertility. Long noncoding RNAs (lncRNAs) play important and wide-ranging roles in the growth and development of plants and have become a major research focus in recent years. Therefore, we explored the characterization and expression change of lncRNAs during ovule development and female gametophytic abortion. RESULTS: In our study, whole-transcriptome strand-specific RNA sequencing (ssRNA-seq) was performed in the ovules of a high-frequency female-sterile rice line (fsv1) and a wild-type rice line (Gui99) at the megaspore mother cell meiosis stage (stage 1), functional megaspore mitosis stage (stage 2) and female gametophyte mature stage (stage 3). By comparing two rice lines, we identified 152, 233, and 197 differentially expressed lncRNAs at the three ovule developmental stages. Functional analysis of the coherent target genes of these differentially expressed lncRNAs indicated that many lncRNAs participate in multiple pathways such as hormone and cellular metabolism and signal transduction. Moreover, there were many differentially expressed lncRNAs acting as the precursors of some miRNAs that are involved in the development of ovules and female gametophytes. In addition, we have found that lncRNAs can act as decoys, competing with mRNAs for binding to miRNAs to maintain the normal expression of genes related to ovule and female gametophyte development. CONCLUSION: These results provide important clues for elucidating the female gametophyte abortion mechanism in rice. This study also expands our understanding about the biological functions of lncRNAs and the annotation of the rice genome.
Assuntos
Regulação da Expressão Gênica de Plantas , Células Germinativas Vegetais/metabolismo , Oryza/genética , Óvulo Vegetal/genética , RNA Longo não Codificante/metabolismo , Perfilação da Expressão Gênica , Genoma de Planta , Sequenciamento de Nucleotídeos em Larga Escala , MicroRNAs/química , MicroRNAs/metabolismo , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Óvulo Vegetal/crescimento & desenvolvimento , Proteínas de Plantas/genética , Precursores de RNA/química , Precursores de RNA/metabolismo , RNA Longo não Codificante/química , RNA Longo não Codificante/fisiologia , RNA Mensageiro/metabolismo , Análise de Sequência de RNARESUMO
Female gametophyte (FG) is crucial for reproduction in flowering plants. Arabidopsis thaliana produces Polygonum-type FGs, which consist of an egg cell, two synergid cells, three antipodal cells and a central cell. Egg cell and central cell are the two female gametes that give rise to the embryo and surrounding endosperm, respectively, after fertilization. During the development of a FG, a single megaspore produced by meiosis undergoes three rounds of mitosis to produce an eight-nucleate cell. A seven-celled FG is formed after cellularization. The central cell initially contains two polar nuclei that fuse during female gametogenesis to form the secondary nucleus. In this study, we developed a gene induction system for analyzing the functions of various genes in developing Arabidopsis FGs. This system allows transgene expression in developing FGs using the heat-inducible Cre-loxP recombination system and FG-specific embryo sac 2 (ES2) promoter. Efficient gene induction was achieved in FGs by incubating flower buds and isolated pistils at 35�C for short periods of time (1-5 min). Gene induction was also induced in developing FGs by heat treatment of isolated ovules using the infrared laser-evoked gene operator (IR-LEGO) system. Expression of a dominant-negative mutant of Sad1/UNC84 (SUN) proteins in developing FGs using the gene induction system developed in this study caused defects in polar nuclear fusion, indicating the roles of SUN proteins in this process. This strategy represents a new tool for analyzing the functions of genes in FG development and FG functions.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Óvulo Vegetal/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Óvulo Vegetal/genéticaRESUMO
Over the last 10 yr there have been major advances in documenting and understanding dynamic changes to DNA methylation, small RNAs, chromatin modifications and chromatin structure that accompany reproductive development in flowering plants, from germline specification to seed maturation. Here I highlight recent advances in the field, mostly made possible by microscopic analysis of epigenetic states or by the ability to isolate specific cell types or tissues and apply omics approaches. I consider in which contexts there is potentially reprogramming vs maintenance or reinforcement of epigenetic states.
Assuntos
Reprogramação Celular/genética , Epigênese Genética , Magnoliopsida/genética , Magnoliopsida/fisiologia , Cromatina/genética , Gametogênese Vegetal/genética , Magnoliopsida/citologia , Reprodução/genéticaAssuntos
RNA Helicases DEAD-box , Sementes , RNA Helicases DEAD-box/metabolismo , RNA Helicases DEAD-box/genética , Sementes/crescimento & desenvolvimento , Sementes/genética , Sementes/fisiologia , Óvulo Vegetal/genética , Óvulo Vegetal/crescimento & desenvolvimento , Óvulo Vegetal/fisiologia , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Fertilidade/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genéticaRESUMO
The development of the female gametophyte (FG) is one of the key processes of life cycle alteration between the haploid gametophyte and the diploid sporophytes in plants and it is required for successful seed development after fertilization. It is well demonstrated that free nuclear mitosis (FNM) of FG is crucial for the development of the ovule. However, studies of the molecular mechanism of ovule and FG development focused mainly on angiosperms, such as Arabidopsis thaliana and further investigation of gymnosperms remains to be completed. Here, Illumina sequencing of six transcriptomic libraries obtained from developing and abortive ovules at different stages during free nuclear mitosis of magagametophyte (FNMM) was used to acquire transcriptome data and gene expression profiles of Pinus tabulaeformis. Six cDNA libraries generated a total of 71.0 million high-quality clean reads that aligned with 63,449 unigenes and the comparison between developing and abortive ovules identified 7174 differentially expressed genes (DEGs). From the functional annotation results, DEGs involved in the cell cycle and phytohormone regulation were highlighted to reveal their biological importance in ovule development. Furthermore, validation of DEGs from the phytohormone signal transduction pathway was performed using quantitative real-time PCR analysis, revealing the dynamics of transcriptional networks and potential key components in the regulation of FG development in P. tabulaeformis were identified. These findings provide new insights into the regulatory mechanisms of ovule development in woody gymnosperms.