RESUMEN
Soil-borne cereal mosaic virus (SBCMV), the causative agent of wheat mosaic, is a Furovirus challenging wheat production all over Europe. Differently from bread wheat, durum wheat shows greater susceptibility and stronger yield penalties, so identification and genetic characterization of resistance sources are major targets for durum genetics and breeding. The Sbm1 locus providing high level of resistance to SBCMV was mapped in bread wheat to the 5DL chromosome arm (Bass in Genome 49:1140-1148, 2006). This excluded the direct use of Sbm1 for durum wheat improvement. Only one major QTL has been mapped in durum wheat, namely QSbm.ubo-2B, on the 2BS chromosome region coincident with Sbm2, already known in bread wheat as reported (Bayles in HGCA Project Report, 2007). Therefore, QSbm.ubo-2B = Sbm2 is considered a pillar for growing durum in SBCMV-affected areas. Herein, we report the fine mapping of Sbm2 based on bi-parental mapping and GWAS, using the Infinium 90 K SNP array and high-throughput KASP®. Fine mapping pointed out a critical haploblock of 3.2 Mb defined by concatenated SNPs successfully converted to high-throughput KASP® markers coded as KUBO. The combination of KUBO-27, wPt-2106-ASO/HRM, KUBO-29, and KUBO-1 allows unequivocal tracing of the Sbm2-resistant haplotype. The interval harbors 52 high- and 41 low-confidence genes, encoding 17 cytochrome p450, three receptor kinases, two defensins, and three NBS-LRR genes. These results pave the way for Sbm2 positional cloning. Importantly, the development of Sbm2 haplotype tagging KASP® provides a valuable case study for improving efficacy of the European variety testing system and, ultimately, the decision-making process related to varietal characterization and choice.
Asunto(s)
Mapeo Cromosómico , Resistencia a la Enfermedad , Enfermedades de las Plantas , Polimorfismo de Nucleótido Simple , Sitios de Carácter Cuantitativo , Triticum , Triticum/genética , Triticum/virología , Enfermedades de las Plantas/virología , Enfermedades de las Plantas/genética , Resistencia a la Enfermedad/genética , Fenotipo , Cromosomas de las Plantas/genética , Virus del Mosaico/patogenicidad , Genes de Plantas , Marcadores GenéticosRESUMEN
Dry yeast extracts (DYE) are applied to vineyards to improve aromatic and secondary metabolic compound content and wine quality; however, systematic information on the underpinning molecular mechanisms is lacking. This work aimed to unravel, through a systematic approach, the metabolic and molecular responses of Sauvignon Blanc berries to DYE treatments. To accomplish this, DYE spraying was performed in a commercial vineyard for two consecutive years. Berries were sampled at several time points after the treatment, and grapes were analyzed for sugars, acidity, free and bound aroma precursors, amino acids, and targeted and untargeted RNA-Seq transcriptional profiles. The results obtained indicated that the DYE treatment did not interfere with the technological ripening parameters of sugars and acidity. Some aroma precursors, including cys-3MH and GSH-3MH, responsible for the typical aromatic nuances of Sauvignon Blanc, were stimulated by the treatment during both vintages. The levels of amino acids and the global RNA-seq transcriptional profiles indicated that DYE spraying upregulated ROS homeostatic and thermotolerance genes, as well as ethylene and jasmonic acid biosynthetic genes, and activated abiotic and biotic stress responses. Overall, the data suggested that the DYE reduced berry oxidative stress through the regulation of specific subsets of metabolic and hormonal pathways.
RESUMEN
In temperate zones, fruit trees regulate their annual growth cycle to seasonal environmental changes. During the cold season, growth is limited by both environmental and genetic factors. After the exposure to low temperature and fulfillment of chilling requirements, mild temperatures promote the growth and flowering. However, an insufficient chilling exposure may lead to nonuniform blooming, with a negative impact on fruit set. To gain insights into flower development in the fruit tree buds, peach is an interesting model, the flower and vegetative bud being distinct organs. To understand how flower bud development is regulated, we integrated cytological observations and epigenetic and chromatin genome-wide data with transcriptional changes to identify the main regulatory factors involved in flower development during chilling accumulation. We demonstrated that growth cessation does not occur in peach flower buds during chilling accumulation, but that there are changes in transcript abundance of key genes of hormone metabolism and flower bud development, distribution of histone modifications (H3K4me3 and H3K27me3) and DNA methylation. Altogether, our findings indicate that during the cold season the flower bud is in a nondormant state and that the chilling experience allows flower differentiation to be completed.
Asunto(s)
Prunus persica , Cromatina/metabolismo , Frío , Flores/fisiología , Regulación de la Expresión Génica de las Plantas , Histonas/metabolismo , Hormonas/metabolismo , Prunus persica/genéticaRESUMEN
Root angle in crops represents a key trait for efficient capture of soil resources. Root angle is determined by competing gravitropic versus antigravitropic offset (AGO) mechanisms. Here we report a root angle regulatory gene termed ENHANCED GRAVITROPISM1 (EGT1) that encodes a putative AGO component, whose loss-of-function enhances root gravitropism. Mutations in barley and wheat EGT1 genes confer a striking root phenotype, where every root class adopts a steeper growth angle. EGT1 encodes an F-box and Tubby domain-containing protein that is highly conserved across plant species. Haplotype analysis found that natural allelic variation at the barley EGT1 locus impacts root angle. Gravitropic assays indicated that Hvegt1 roots bend more rapidly than wild-type. Transcript profiling revealed Hvegt1 roots deregulate reactive oxygen species (ROS) homeostasis and cell wall-loosening enzymes and cofactors. ROS imaging shows that Hvegt1 root basal meristem and elongation zone tissues have reduced levels. Atomic force microscopy measurements detected elongating Hvegt1 root cortical cell walls are significantly less stiff than wild-type. In situ analysis identified HvEGT1 is expressed in elongating cortical and stele tissues, which are distinct from known root gravitropic perception and response tissues in the columella and epidermis, respectively. We propose that EGT1 controls root angle by regulating cell wall stiffness in elongating root cortical tissue, counteracting the gravitropic machinery's known ability to bend the root via its outermost tissues. We conclude that root angle is controlled by EGT1 in cereal crops employing an antigravitropic mechanism.
Asunto(s)
Productos Agrícolas , Gravitropismo , Hordeum , Proteínas de Plantas , Raíces de Plantas , Pared Celular/química , Productos Agrícolas/química , Productos Agrícolas/genética , Productos Agrícolas/crecimiento & desarrollo , Gravitropismo/genética , Hordeum/química , Hordeum/genética , Hordeum/crecimiento & desarrollo , Microscopía de Fuerza Atómica , Proteínas de Plantas/genética , Proteínas de Plantas/fisiología , Raíces de Plantas/química , Raíces de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo , Especies Reactivas de Oxígeno/metabolismo , Transcripción GenéticaRESUMEN
BACKGROUND: Perennial fruit trees display a growth behaviour characterized by annual cycling between growth and dormancy, with complex physiological features. Rosaceae fruit trees represent excellent models for studying not only the fruit growth/patterning but also the progression of the reproductive cycle depending upon the impact of climate conditions. Additionally, current developments in high-throughput technologies have impacted Rosaceae tree research while investigating genome structure and function as well as (epi)genetic mechanisms involved in important developmental and environmental response processes during fruit tree growth. Among epigenetic mechanisms, chromatin remodelling mediated by histone modifications and other chromatin-related processes play a crucial role in gene modulation, controlling gene expression. Chromatin immunoprecipitation is an effective technique to investigate chromatin dynamics in plants. This technique is generally applied for studies on chromatin states and enrichment of post-transcriptional modifications (PTMs) in histone proteins. RESULTS: Peach is considered a model organism among climacteric fruits in the Rosaceae family for studies on bud formation, dormancy, and organ differentiation. In our work, we have primarily established specific protocols for chromatin extraction and immunoprecipitation in reproductive tissues of peach (Prunus persica). Subsequently, we focused our investigations on the role of two chromatin marks, namely the trimethylation of histone H3 at lysine in position 4 (H3K4me3) and trimethylation of histone H3 at lysine 27 (H3K27me3) in modulating specific gene expression. Bud dormancy and fruit growth were investigated in a nectarine genotype called Fantasia as our model system. CONCLUSIONS: We present general strategies to optimize ChIP protocols for buds and mesocarp tissues of peach and analyze the correlation between gene expression and chromatin mark enrichment/depletion. The procedures proposed may be useful to evaluate any involvement of histone modifications in the regulation of gene expression during bud dormancy progression and core ripening in fruits.
RESUMEN
Disease lesion mimic (DLM) or necrotic mutants display necrotic lesions in the absence of pathogen infections. They can show improved resistance to some pathogens and their molecular dissection can contribute to revealing components of plant defense pathways. Although forward-genetics strategies to find genes causal to mutant phenotypes are available in crops, these strategies require the production of experimental cross populations, mutagenesis, or gene editing and are time- and resource-consuming or may have to deal with regulated plant materials. In this study, we described a collection of 34 DLM mutants in barley (Hordeum vulgare L.) and applied a novel method called complementation by sequencing (CBS), which enables the identification of the gene responsible for a mutant phenotype given the availability of two or more chemically mutagenized individuals showing the same phenotype. Complementation by sequencing relies on the feasibility to obtain all induced mutations present in chemical mutants and on the low probability that different individuals share the same mutated genes. By CBS, we identified a cytochrome P450 CYP71P1 gene as responsible for orange blotch DLM mutants, including the historical barley nec3 locus. By comparative phylogenetic analysis we showed that CYP71P1 gene family emerged early in angiosperm evolution but has been recurrently lost in some lineages including Arabidopsis thaliana (L.) Heynh. Complementation by sequencing is a straightforward cost-effective approach to clone genes controlling phenotypes in a chemically mutagenized collection. The TILLMore (TM) collection will be instrumental for understanding the molecular basis of DLM phenotypes and to contribute knowledge about mechanisms of host-pathogen interaction.
Asunto(s)
Arabidopsis , Hordeum , Arabidopsis/genética , Clonación Molecular , Genes de Plantas , Hordeum/genética , Mutación , FilogeniaRESUMEN
Osmotic adjustment (OA) is a major component of drought resistance in crops. The genetic basis of OA in wheat and other crops remains largely unknown. In this study, 248 field-grown durum wheat elite accessions grown under well-watered conditions, underwent a progressively severe drought treatment started at heading. Leaf samples were collected at heading and 17 days later. The following traits were considered: flowering time (FT), leaf relative water content (RWC), osmotic potential (ψs), OA, chlorophyll content (SPAD), and leaf rolling (LR). The high variability (3.89-fold) in OA among drought-stressed accessions resulted in high repeatability of the trait (h2 = 72.3%). Notably, a high positive correlation (r = 0.78) between OA and RWC was found under severe drought conditions. A genome-wide association study (GWAS) revealed 15 significant QTLs (Quantitative Trait Loci) for OA (global R2 = 63.6%), as well as eight major QTL hotspots/clusters on chromosome arms 1BL, 2BL, 4AL, 5AL, 6AL, 6BL, and 7BS, where a higher OA capacity was positively associated with RWC and/or SPAD, and negatively with LR, indicating a beneficial effect of OA on the water status of the plant. The comparative analysis with the results of 15 previous field trials conducted under varying water regimes showed concurrent effects of five OA QTL cluster hotspots on normalized difference vegetation index (NDVI), thousand-kernel weight (TKW), and/or grain yield (GY). Gene content analysis of the cluster regions revealed the presence of several candidate genes, including bidirectional sugar transporter SWEET, rhomboid-like protein, and S-adenosyl-L-methionine-dependent methyltransferases superfamily protein, as well as DREB1. Our results support OA as a valuable proxy for marker-assisted selection (MAS) aimed at enhancing drought resistance in wheat.
Asunto(s)
Estudio de Asociación del Genoma Completo , Triticum , Sequías , Sitios de Carácter Cuantitativo , Triticum/genética , AguaRESUMEN
Superficial scald is a post-harvest chilling storage injury leading to browning of the surface of the susceptible cv Granny Smith apples. Wounding of skins has been reported to play a preventive role on scald development however its underlying molecular factors are unknown. We have artificially wounded the epidermal and sub-epidermal layers of apple skins consistently obtaining the prevention of superficial scald in the surroundings of the wounds during two independent vintages. Time course RNA-Seq analyses of the transcriptional changes in wounded versus unwounded skins revealed that two transcriptional waves occurred. An early wave included genes up-regulated by wounding already after 6 h, highlighting a specific transcriptional rearrangement of genes connected to the biosynthesis and signalling of JA, ethylene and ABA. A later transcriptional wave, occurring after three months of cold storage, included genes up-regulated exclusively in unwounded skins and was prevented from its occurrence in wounded skins. A significant portion of these genes was related to decay of tissues and to the senescence hormones ABA, JA and ethylene. Such changes suggest a wound-inducible reversed hormonal balance during post-harvest storage which may explain the local inhibition of scald in wounded tissues, an aspect that will need further studies for its mechanistic explanation.
Asunto(s)
Conservación de Alimentos , Frutas , Regulación de la Expresión Génica de las Plantas , Malus , RNA-Seq , Frutas/genética , Frutas/metabolismo , Malus/genética , Malus/metabolismoRESUMEN
Maize root responds to nitrate by modulating its development through the coordinated action of many interacting players. Nitric oxide is produced in primary root early after the nitrate provision, thus inducing root elongation. In this study, RNA sequencing was applied to discover the main molecular signatures distinguishing the response of maize root to nitrate according to their dependency on, or independency of, nitric oxide, thus discriminating the signaling pathways regulated by nitrate through nitric oxide from those regulated by nitrate itself of by further downstream factors. A set of subsequent detailed functional annotation tools (Gene Ontology enrichment, MapMan, KEGG reconstruction pathway, transcription factors detection) were used to gain further information and the lateral root density was measured both in the presence of nitrate and in the presence of nitrate plus cPTIO, a specific NO scavenger, and compared to that observed for N-depleted roots. Our results led us to identify six clusters of transcripts according to their responsiveness to nitric oxide and to their regulation by nitrate provision. In general, shared and specific features for the six clusters were identified, allowing us to determine the overall root response to nitrate according to its dependency on nitric oxide.
Asunto(s)
Regulación de la Expresión Génica de las Plantas , Nitratos/metabolismo , Raíces de Plantas/metabolismo , Transcriptoma , Zea mays/metabolismo , Benzoatos , Fertilizantes , Imidazoles , Óxido Nítrico/metabolismoRESUMEN
The root growth angle defines how roots grow toward the gravity vector and is among the most important determinants of root system architecture. It controls water uptake capacity, nutrient use efficiency, stress resilience, and, as a consequence, yield of crop plants. We demonstrated that the egt2 (enhanced gravitropism 2) mutant of barley exhibits steeper root growth of seminal and lateral roots and an auxin-independent higher responsiveness to gravity compared to wild-type plants. We cloned the EGT2 gene by a combination of bulked-segregant analysis and whole genome sequencing. Subsequent validation experiments by an independent CRISPR/Cas9 mutant allele demonstrated that egt2 encodes a STERILE ALPHA MOTIF domain-containing protein. In situ hybridization experiments illustrated that EGT2 is expressed from the root cap to the elongation zone. We demonstrated the evolutionary conserved role of EGT2 in root growth angle control between barley and wheat by knocking out the EGT2 orthologs in the A and B genomes of tetraploid durum wheat. By combining laser capture microdissection with RNA sequencing, we observed that seven expansin genes were transcriptionally down-regulated in the elongation zone. This is consistent with a role of EGT2 in this region of the root where the effect of gravity sensing is executed by differential cell elongation. Our findings suggest that EGT2 is an evolutionary conserved regulator of root growth angle in barley and wheat that could be a valuable target for root-based crop improvement strategies in cereals.
Asunto(s)
Gravitropismo , Hordeum/fisiología , Proteínas de Plantas/fisiología , Raíces de Plantas/crecimiento & desarrollo , Motivo alfa Estéril , Triticum/fisiología , Pared Celular/metabolismo , Secuencia Conservada , Evolución Molecular , Técnicas de Inactivación de Genes , Genes de Plantas , Hordeum/genética , Hordeum/crecimiento & desarrollo , Ácidos Indolacéticos/metabolismo , Mutación , Proteínas de Plantas/química , Proteínas de Plantas/genética , Triticum/genética , Triticum/crecimiento & desarrolloRESUMEN
In deciduous fruit trees, entrance into dormancy occurs in later summer/fall, concomitantly with the shortening of day length and decrease in temperature. Dormancy can be divided into endodormancy, ecodormancy and paradormancy. In Prunus species flower buds, entrance into the dormant stage occurs when the apical meristem is partially differentiated; during dormancy, flower verticils continue their growth and differentiation. Each species and/or cultivar requires exposure to low winter temperature followed by warm temperatures, quantified as chilling and heat requirements, to remove the physiological blocks that inhibit budburst. A comprehensive meta-analysis of transcriptomic studies on flower buds of sweet cherry, apricot and peach was conducted, by investigating the gene expression profiles during bud endo- to ecodormancy transition in genotypes differing in chilling requirements. Conserved and distinctive expression patterns were observed, allowing the identification of gene specifically associated with endodormancy or ecodormancy. In addition to the MADS-box transcription factor family, hormone-related genes, chromatin modifiers, macro- and micro-gametogenesis related genes and environmental integrators, were identified as novel biomarker candidates for flower bud development during winter in stone fruits. In parallel, flower bud differentiation processes were associated to dormancy progression and termination and to environmental factors triggering dormancy phase-specific gene expression.
Asunto(s)
Flores/crecimiento & desarrollo , Genes de Plantas , Prunus/genética , ARN de Planta/biosíntesis , Transcriptoma , Epigénesis Genética , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Proteínas de Dominio MADS/biosíntesis , Proteínas de Dominio MADS/genética , Óvulo Vegetal/fisiología , Filogenia , Reguladores del Crecimiento de las Plantas/fisiología , Proteínas de Plantas/biosíntesis , Proteínas de Plantas/genética , Polen/fisiología , Prunus/crecimiento & desarrollo , Prunus/efectos de la radiación , Prunus armeniaca/genética , Prunus armeniaca/crecimiento & desarrollo , Prunus armeniaca/efectos de la radiación , Prunus avium/genética , Prunus avium/crecimiento & desarrollo , Prunus avium/efectos de la radiación , Prunus persica/genética , Prunus persica/crecimiento & desarrollo , Prunus persica/efectos de la radiación , ARN de Planta/genética , RNA-Seq , Estaciones del Año , Especificidad de la Especie , Luz Solar , Temperatura , Factores de Transcripción/biosíntesis , Factores de Transcripción/genéticaRESUMEN
Consumers' choices are mainly based on fruit external characteristics such as the final size, weight, and shape. The majority of edible fruit are by tree fruit species, among which peach is the genomic and genetic reference for Prunus. In this research, we used a peach with a slow ripening (SR) phenotype, identified in the Fantasia (FAN) nectarine, associated with misregulation of genes involved in mesocarp identity and showing a reduction of final fruit size. By investigating the ploidy level, we observed a progressive increase in endoreduplication in mesocarp, which occurred in the late phases of FAN fruit development, but not in SR fruit. During fruit growth, we also detected that genes involved in endoreduplication were differentially modulated in FAN compared to SR. The differential transcriptional outputs were consistent with different chromatin states at loci of endoreduplication genes. The impaired expression of genes controlling cell cycle and endocycle as well as those claimed to play a role in fruit tissue identity result in the small final size of SR fruit.
Asunto(s)
Perfilación de la Expresión Génica/métodos , Prunus persica/fisiología , Sitios de Carácter Cuantitativo , Ciclo Celular , Endorreduplicación , Regulación de la Expresión Génica de las Plantas , Fenotipo , Proteínas de Plantas/genética , Ploidias , Prunus persica/genética , Análisis de Secuencia de ARNRESUMEN
In the coming decades, larger genetic gains in yield will be necessary to meet projected demand, and this must be achieved despite the destabilizing impacts of climate change on crop production. The root systems of crops capture the water and nutrients needed to support crop growth, and improved root systems tailored to the challenges of specific agricultural environments could improve climate resiliency. Each component of root initiation, growth and development is controlled genetically and responds to the environment, which translates to a complex quantitative system to navigate for the breeder, but also a world of opportunity given the right tools. In this review, we argue that it is important to know more about the 'hidden half' of crop plants and hypothesize that crop improvement could be further enhanced using approaches that directly target selection for root system architecture. To explore these issues, we focus predominantly on bread wheat (Triticum aestivum L.), a staple crop that plays a major role in underpinning global food security. We review the tools available for root phenotyping under controlled and field conditions and the use of these platforms alongside modern genetics and genomics resources to dissect the genetic architecture controlling the wheat root system. To contextualize these advances for applied wheat breeding, we explore questions surrounding which root system architectures should be selected for, which agricultural environments and genetic trait configurations of breeding populations are these best suited to, and how might direct selection for these root ideotypes be implemented in practice.
Asunto(s)
Cambio Climático , Fitomejoramiento , Raíces de Plantas/fisiología , Triticum/genética , Productos Agrícolas/genética , Genes de Plantas , Fenotipo , Raíces de Plantas/genética , Triticum/fisiologíaRESUMEN
In maize, nitrate regulates root development thanks to the coordinated action of many players. In this study, the involvement of strigolactones (SLs) and auxin as putative components of the nitrate regulation of lateral root (LR) was investigated. To this aim, the endogenous SL content of maize root in response to nitrate was assessed by liquid chromatography with tandem mass Spectrometry (LC-MS/MS) and measurements of LR density in the presence of analogues or inhibitors of auxin and SLs were performed. Furthermore, an untargeted RNA-sequencing (RNA-seq)-based approach was used to better characterize the participation of auxin and SLs to the transcriptional signature of maize root response to nitrate. Our results suggested that N deprivation induces zealactone and carlactonoic acid biosynthesis in root, to a higher extent if compared to P-deprived roots. Moreover, data on LR density led to hypothesize that the induction of LR development early occurring upon nitrate supply involves the inhibition of SL biosynthesis, but that the downstream target of SL shutdown, besides auxin, also includes additional unknown players. Furthermore, RNA-seq results provided a set of putative markers for the auxin- or SL-dependent action of nitrate, meanwhile also allowing to identify novel components of the molecular regulation of maize root response to nitrate. Globally, the existence of at least four different pathways was hypothesized: one dependent on auxin, a second one mediated by SLs, a third deriving from the SL-auxin interplay, and a last one attributable to nitrate itself through further downstream signals. Further work will be necessary to better assess the reliability of the model proposed.
Asunto(s)
Compuestos Heterocíclicos con 3 Anillos/metabolismo , Ácidos Indolacéticos/metabolismo , Lactonas/metabolismo , Nitratos/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Zea mays/crecimiento & desarrollo , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Germinación , Hexanonas/farmacología , Nitratos/farmacología , Nitrógeno/metabolismo , Orobanchaceae/metabolismo , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/metabolismo , Análisis de Secuencia de ARN , Espectrometría de Masas en Tándem , Triazoles/farmacología , Zea mays/efectos de los fármacos , Zea mays/metabolismoRESUMEN
Nitrogen (N) is an essential macronutrient for crops. Plants have developed several responses to N fluctuations, thus optimizing the root architecture in response to N availability. Nitrate and ammonium are the main inorganic N forms taken up by plants, and act as both nutrients and signals, affecting gene expression and plant development. In this study, RNA-sequencing was applied to gain comprehensive information on the pathways underlying the response of maize root, pre-treated in an N-deprived solution, to the provision of nitrate or ammonium. The analysis of the transcriptome shows that nitrate and ammonium regulate overlapping and distinct pathways, thus leading to different responses. Ammonium activates the response to stress, while nitrate acts as a negative regulator of transmembrane transport. Both the N-source repress genes related to the cytoskeleton and reactive oxygen species detoxification. Moreover, the presence of ammonium induces the accumulation of anthocyanins, while also reducing biomass and chlorophyll and flavonoids accumulation. Furthermore, the later physiological effects of these nutrients were evaluated through the assessment of shoot and root growth, leaf pigment content and the amino acid concentrations in root and shoot, confirming the existence of common and distinct features in response to the two nitrogen forms.
Asunto(s)
Compuestos de Amonio/farmacología , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Nitratos/farmacología , Nitrógeno/metabolismo , Transcriptoma , Zea mays/fisiología , Aminoácidos , Análisis por Conglomerados , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Raíces de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/fisiología , Análisis de Secuencia de ARN , Zea mays/genética , Zea mays/crecimiento & desarrolloRESUMEN
During their lifespan, plants respond to a multitude of stressful factors. Dynamic changes in chromatin and concomitant transcriptional variations control stress response and adaptation, with epigenetic memory mechanisms integrating environmental conditions and appropriate developmental programs over the time. Here we analyzed transcriptome and genome-wide histone modifications of maize plants subjected to a mild and prolonged drought stress just before the flowering transition. Stress was followed by a complete recovery period to evaluate drought memory mechanisms. Three categories of stress-memory genes were identified: i) "transcriptional memory" genes, with stable transcriptional changes persisting after the recovery; ii) "epigenetic memory candidate" genes in which stress-induced chromatin changes persist longer than the stimulus, in absence of transcriptional changes; iii) "delayed memory" genes, not immediately affected by the stress, but perceiving and storing stress signal for a delayed response. This last memory mechanism is described for the first time in drought response. In addition, applied drought stress altered floral patterning, possibly by affecting expression and chromatin of flowering regulatory genes. Altogether, we provided a genome-wide map of the coordination between genes and chromatin marks utilized by plants to adapt to a stressful environment, describing how this serves as a backbone for setting stress memory.
Asunto(s)
Aclimatación , Adaptación Fisiológica/genética , Epigénesis Genética , Flores/fisiología , Estrés Fisiológico/genética , Zea mays/fisiología , Cromatina/metabolismo , Mapeo Cromosómico , Cromosomas de las Plantas/fisiología , Sequías , Epigenómica , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Ontología de Genes , Código de Histonas , Histonas/genética , Histonas/metabolismo , Inmunoprecipitación , Desarrollo de la Planta/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Análisis de Componente Principal , Análisis de Secuencia de ARN , TranscriptomaRESUMEN
Histone deacetylases (HDACs) catalyze the removal of acetyl groups from acetylated histone tails that consequently interact more closely with DNA, leading to chromatin state refractory to transcription. Zea mays HDA108 belongs to the Rpd3/HDA1 HDAC family and is ubiquitously expressed during development. The newly isolated hda108/hda108 insertional mutant exhibited many developmental defects: significant reduction in plant height, alterations of shoot and leaf development, and alterations of inflorescence patterning and fertility. Western blot analyses and immunolocalization experiments revealed an evident increase in histone acetylation, accompanied by a marked reduction in H3K9 dimethylation, in mutant nuclei. The DNA methylation status, in the CHG sequence context, and the transcript level of ribosomal sequences were also affected in hda108 mutants, while enrichment in H3 and H4 acetylation characterizes both repetitive and nonrepetitive transcriptional up-regulated loci. RNA-Seq of both young leaf and anthers indicated that transcription factor expression is highly affected and that the pollen developmental program is disrupted in hda108 mutants. Crosses between hda108/hda108 and epiregulator mutants did not produce any double mutant progeny indicating possible genetic interactions of HDA108 with distinct epigenetic pathways. Our findings indicate that HDA108 is directly involved in regulation of maize development, fertility, and epigenetic regulation of genome activity.
Asunto(s)
Silenciador del Gen , Histona Desacetilasas/metabolismo , ARN Ribosómico/genética , Reproducción , Zea mays/fisiología , Acetilación , Biología Computacional/métodos , Metilación de ADN , Epigénesis Genética , Técnicas de Inactivación de Genes , Ontología de Genes , Sitios Genéticos , Histonas/metabolismo , Mutación , Fenotipo , Procesamiento Proteico-PostraduccionalRESUMEN
Non-coding RNA transcripts, such as long non-coding RNAs, miRNAs, siRNAs, and transposon-originating transcripts, are involved in the regulation of RNA stability, protein translation, and/or the modulation of chromatin states. RNA-Seq can be used to catalog this diversity of novel transcripts and a joint analysis of these transcriptomic data can provide useful insights into epigenetic regulation of dynamic responses such as the stress response, which may not be deciphered from individual analysis of single transcript categories. Here, we present a protocol that allows the identification and analysis of small RNAs and long non-coding RNAs, together with the comparison of these species between different sample types.
Asunto(s)
Secuenciación de Nucleótidos de Alto Rendimiento/métodos , ARN Largo no Codificante/genética , ARN Pequeño no Traducido/genética , Análisis de Secuencia de ARN/métodos , Zea mays/genética , Elementos Transponibles de ADN , Epigénesis Genética , Perfilación de la Expresión Génica/métodos , Proteínas de Plantas/genética , ARN de Planta/genéticaRESUMEN
BACKGROUND: RNA-directed DNA methylation (RdDM) is a plant-specific epigenetic process that relies on the RNA polymerase IV (Pol IV) for the production of 24 nucleotide small interfering RNAs (siRNA) that guide the cytosine methylation and silencing of genes and transposons. Zea mays RPD1/RMR6 gene encodes the largest subunit of Pol IV and is required for normal plant development, paramutation, transcriptional repression of certain transposable elements (TEs) and transcriptional regulation of specific alleles. RESULTS: In this study we applied a total RNA-Seq approach to compare the B73 and rpd1/rmr6 leaf transcriptomes. Although previous studies indicated that loss of siRNAs production in RdDM mutants provokes a strong loss of CHH DNA methylation but not massive gene or TEs transcriptional activation in both Arabidopsis and maize, our total RNA-Seq analysis of rpd1/rmr6 transcriptome reveals that loss of Pol IV activity causes a global increase in the transcribed fraction of the maize genome. Our results point to the genes with nearby TE insertions as being the most strongly affected by Pol IV-mediated gene silencing. TEs modulation of nearby gene expression is linked to alternative methylation profiles on gene flanking regions, and these profiles are strictly dependent on specific characteristics of the TE member inserted. Although Pol IV is essential for the biogenesis of siRNAs, the genes with associated siRNA loci are less affected by the pol IV mutation. CONCLUSIONS: This deep and integrated analysis of gene expression, TEs distribution, smallRNA targeting and DNA methylation levels, reveals that loss of Pol IV activity globally affects genome regulation, pointing at TEs as modulator of nearby gene expression and indicating the existence of multiple level epigenetic silencing mechanisms. Our results also suggest a predominant role of the Pol IV-mediated RdDM pathway in genome dominance regulation, and subgenome stability and evolution in maize.
Asunto(s)
Elementos Transponibles de ADN , ARN Polimerasas Dirigidas por ADN/metabolismo , Regulación de la Expresión Génica de las Plantas , Genoma de Planta , Zea mays/enzimología , Zea mays/genética , Metilación de ADN , ADN de Plantas/metabolismo , Mutación , Hojas de la Planta/metabolismo , ARN de Planta , ARN Interferente Pequeño , Análisis de Secuencia de ARN , TranscriptomaRESUMEN
Plant's response and adaptation to abiotic stresses involve sophisticated genetic and epigenetic regulatory systems. To obtain a global view of molecular response to osmotic stresses, including the non-coding portion of genome, we conducted a total leaf transcriptome analysis on maize plants subjected to prolonged drought and salt stresses. Stress application to both B73 wild type and the epiregulator mutant rpd1-1/rmr6 allowed dissection of the epigenetic component of stress response. Coupling total RNA-Seq and transcriptome re-assembly we annotated thousands of new maize transcripts, together with 13,387 lncRNAs that may play critical roles in regulating gene expression. Differential expression analysis revealed hundreds of genes modulated by long-term stress application, including also many lncRNAs and transposons specifically induced by stresses. The amplitude and dynamic of the stress-modulated gene sets are very different between B73 and rpd1-1/rmr6 mutant plants, as result of stress-like effect on genome regulation caused by the mutation itself, which activates many stress-related genes even in control condition. The analyzed extensive set of total RNA-Seq data, together with the improvement of the transcriptome and the identification of the non-coding portion of the transcriptome give a revealing insight into the genetic and epigenetic mechanism responsible for maize molecular response to abiotic stresses.