RESUMEN
Chloroplast biogenesis is dependent on master regulators from the GOLDEN2-LIKE (GLK) family of transcription factors. However, glk mutants contain residual chlorophyll, indicating that other proteins must be involved. Here, we identify MYB-related transcription factors as regulators of chloroplast biogenesis in the liverwort Marchantia polymorpha and angiosperm Arabidopsis thaliana. In both species, double-mutant alleles in MYB-related genes show very limited chloroplast development, and photosynthesis gene expression is perturbed to a greater extent than in GLK mutants. Genes encoding enzymes of chlorophyll biosynthesis are controlled by MYB-related and GLK proteins, whereas those allowing CO2 fixation, photorespiration, and photosystem assembly and repair require MYB-related proteins. Regulation between the MYB-related and GLK transcription factors appears more extensive in A. thaliana than in M. polymorpha. Thus, MYB-related and GLK genes have overlapping as well as distinct targets. We conclude that MYB-related and GLK transcription factors orchestrate chloroplast development in land plants.
RESUMEN
Grafting is possible in both animals and plants. Although in animals the process requires surgery and is often associated with rejection of non-self, in plants grafting is widespread, and has been used since antiquity for crop improvement1. However, in the monocotyledons, which represent the second largest group of terrestrial plants and include many staple crops, the absence of vascular cambium is thought to preclude grafting2. Here we show that the embryonic hypocotyl allows intra- and inter-specific grafting in all three monocotyledon groups: the commelinids, lilioids and alismatids. We show functional graft unions through histology, application of exogenous fluorescent dyes, complementation assays for movement of endogenous hormones, and growth of plants to maturity. Expression profiling identifies genes that unify the molecular response associated with grafting in monocotyledons and dicotyledons, but also gene families that have not previously been associated with tissue union. Fusion of susceptible wheat scions to oat rootstocks confers resistance to the soil-borne pathogen Gaeumannomyces graminis. Collectively, these data overturn the consensus that monocotyledons cannot form graft unions, and identify the hypocotyl (mesocotyl in grasses) as a meristematic tissue that allows this process. We conclude that graft compatibility is a shared ability among seed-bearing plants.
Asunto(s)
Avena , Raíces de Plantas , Brotes de la Planta , Trasplantes , Triticum , Ascomicetos/patogenicidad , Avena/embriología , Avena/microbiología , Hipocótilo , Meristema , Raíces de Plantas/embriología , Raíces de Plantas/microbiología , Brotes de la Planta/embriología , Brotes de la Planta/microbiología , Triticum/embriología , Triticum/microbiologíaRESUMEN
Gynandropsis gynandra (Cleomaceae) is a cosmopolitan leafy vegetable and medicinal plant, which has also been used as a model to study C4 photosynthesis due to its evolutionary proximity to C3 Arabidopsis (Arabidopsis thaliana). Here, we present the genome sequence of G. gynandra, anchored onto 17 main pseudomolecules with a total length of 740 Mb, an N50 of 42 Mb and 30,933 well-supported gene models. The G. gynandra genome and previously released genomes of C3 relatives in the Cleomaceae and Brassicaceae make an excellent model for studying the role of genome evolution in the transition from C3 to C4 photosynthesis. Our analyses revealed that G. gynandra and its C3 relative Tarenaya hassleriana shared a whole-genome duplication event (Gg-α), then an addition of a third genome (Th-α, +1×) took place in T. hassleriana but not in G. gynandra. Analysis of syntenic copy number of C4 photosynthesis-related gene families indicates that G. gynandra generally retained more duplicated copies of these genes than C3T. hassleriana, and also that the G. gynandra C4 genes might have been under positive selection pressure. Both whole-genome and single-gene duplication were found to contribute to the expansion of the aforementioned gene families in G. gynandra. Collectively, this study enhances our understanding of the polyploidy history, gene duplication and retention, as well as their impact on the evolution of C4 photosynthesis in Cleomaceae.
Asunto(s)
Arabidopsis , Brassicaceae , Magnoliopsida , Duplicación de Gen , Magnoliopsida/genética , Brassicaceae/genética , Arabidopsis/genética , Fotosíntesis/genética , Evolución MolecularRESUMEN
Model species continue to underpin groundbreaking plant science research. At the same time, the phylogenetic resolution of the land plant Tree of Life continues to improve. The intersection of these two research paths creates a unique opportunity to further extend the usefulness of model species across larger taxonomic groups. Here we promote the utility of the Arabidopsis thaliana model species, especially the ability to connect its genetic and functional resources, to species across the entire Brassicales order. We focus on the utility of using genomics and phylogenomics to bridge the evolution and diversification of several traits across the Brassicales to the resources in Arabidopsis, thereby extending scope from a model species by establishing a "model clade". These Brassicales-wide traits are discussed in the context of both the model species Arabidopsis thaliana and the family Brassicaceae. We promote the utility of such a "model clade" and make suggestions for building global networks to support future studies in the model order Brassicales.
RESUMEN
Land plants comprise two large monophyletic lineages, the vascular plants and the bryophytes, which diverged from their most recent common ancestor approximately 480 million years ago. Of the three lineages of bryophytes, only the mosses and the liverworts are systematically investigated, while the hornworts are understudied. Despite their importance for understanding fundamental questions of land plant evolution, they only recently became amenable to experimental investigation, with Anthoceros agrestis being developed as a hornwort model system. Availability of a high-quality genome assembly and a recently developed genetic transformation technique makes A. agrestis an attractive model species for hornworts. Here we describe an updated and optimized transformation protocol for A. agrestis, which can be successfully used to genetically modify one more strain of A. agrestis and three more hornwort species, Anthoceros punctatus, Leiosporoceros dussii, and Phaeoceros carolinianus. The new transformation method is less laborious, faster, and results in the generation of greatly increased numbers of transformants compared with the previous method. We have also developed a new selection marker for transformation. Finally, we report the development of a set of different cellular localization signal peptides for hornworts providing new tools to better understand the hornwort cell biology.
Asunto(s)
Anthocerotophyta , Briófitas , Embryophyta , Anthocerotophyta/genética , Filogenia , Briófitas/genética , SemillasRESUMEN
In leaves of C4 plants, the reactions of photosynthesis become restricted between two compartments. Typically, this allows accumulation of C4 acids in mesophyll (M) cells and subsequent decarboxylation in the bundle sheath (BS). In C4 grasses, proliferation of plasmodesmata between these cell types is thought to increase cell-to-cell connectivity to allow efficient metabolite movement. However, it is not known whether C4 dicotyledons also show this enhanced plasmodesmal connectivity and so whether this is a general requirement for C4 photosynthesis is not clear. How M and BS cells in C4 leaves become highly connected is also not known. We investigated these questions using 3D- and 2D-electron microscopy on the C4 dicotyledon Gynandropsis gynandra as well as phylogenetically close C3 relatives. The M-BS interface of C4 G. gynandra showed higher plasmodesmal frequency compared with closely related C3 species. Formation of these plasmodesmata was induced by light. Pharmacological agents that perturbed photosynthesis reduced the number of plasmodesmata, but this inhibitory effect could be reversed by the provision of exogenous sucrose. We conclude that enhanced formation of plasmodesmata between M and BS cells is wired to the induction of photosynthesis in C4 G. gynandra.
Asunto(s)
Magnoliopsida , Células del Mesófilo , Células del Mesófilo/metabolismo , Plasmodesmos/metabolismo , Hojas de la Planta/metabolismo , Fotosíntesis , PoaceaeRESUMEN
Common purslane (Portulaca oleracea) integrates both C4 and crassulacean acid metabolism (CAM) photosynthesis pathways and is a promising model plant to explore C4-CAM plasticity. Here, we report a high-quality chromosome-level genome of nicotinamide adenine dinucleotide (NAD)-malic enzyme (ME) subtype common purslane that provides evidence for 2 rounds of whole-genome duplication (WGD) with an ancient WGD (P-ß) in the common ancestor to Portulacaceae and Cactaceae around 66.30 million years ago (Mya) and another (Po-α) specific to common purslane lineage around 7.74 Mya. A larger number of gene copies encoding key enzymes/transporters involved in C4 and CAM pathways were detected in common purslane than in related species. Phylogeny, conserved functional site, and collinearity analyses revealed that the Po-α WGD produced the phosphoenolpyruvate carboxylase-encoded gene copies used for photosynthesis in common purslane, while the P-ß WGD event produced 2 ancestral genes of functionally differentiated (C4- and CAM-specific) beta carbonic anhydrases involved in the C4 + CAM pathways. Additionally, cis-element enrichment analysis in the promoters showed that CAM-specific genes have recruited both evening and midnight circadian elements as well as the Abscisic acid (ABA)-independent regulatory module mediated by ethylene-response factor cis-elements. Overall, this study provides insights into the origin and evolutionary process of C4 and CAM pathways in common purslane, as well as potential targets for engineering crops by integrating C4 or CAM metabolism.
Asunto(s)
Portulaca , Portulaca/genética , Portulaca/metabolismo , Duplicación de Gen , Metabolismo Ácido de las Crasuláceas , Evolución Biológica , Filogenia , Fotosíntesis/genéticaRESUMEN
Leaves of shade-avoiding plants such as Arabidopsis (Arabidopsis thaliana) change their growth pattern and position in response to low red to far-red ratios (LRFRs) encountered in dense plant communities. Under LRFR, transcription factors of the phytochrome-interacting factor (PIF) family are derepressed. PIFs induce auxin production, which is required for promoting leaf hyponasty, thereby favoring access to unfiltered sunlight. Abscisic acid (ABA) has also been implicated in the control of leaf hyponasty, with gene expression patterns suggesting that LRFR regulates the ABA response. Here, we show that LRFR leads to a rapid increase in ABA levels in leaves. Changes in ABA levels depend on PIFs, which regulate the expression of genes encoding isoforms of the enzyme catalyzing a rate-limiting step in ABA biosynthesis. Interestingly, ABA biosynthesis and signaling mutants have more erect leaves than wild-type Arabidopsis under white light but respond less to LRFR. Consistent with this, ABA application decreases leaf angle under white light; however, this response is inhibited under LRFR. Tissue-specific interference with ABA signaling indicates that an ABA response is required in different cell types for LRFR-induced hyponasty. Collectively, our data indicate that LRFR triggers rapid PIF-mediated ABA production. ABA plays a different role in controlling hyponasty under white light than under LRFR. Moreover, ABA exerts its activity in multiple cell types to control leaf position.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Fitocromo , Arabidopsis/metabolismo , Ácido Abscísico/farmacología , Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Fitocromo/metabolismo , Regulación de la Expresión Génica de las PlantasRESUMEN
Compared with the ancestral C3 state, C4 photosynthesis occurs at higher rates with improved water and nitrogen use efficiencies. In both C3 and C4 plants, rates of photosynthesis increase with light intensity and are maximal around midday. We determined that in the absence of light or temperature fluctuations, photosynthesis in maize (Zea mays) peaks in the middle of the subjective photoperiod. To investigate the molecular processes associated with these temporal changes, we performed RNA sequencing of maize mesophyll and bundle sheath strands over a 24-h time course. Preferential expression of C4 cycle genes in these cell types was strongest between 6 and 10 h after dawn when rates of photosynthesis were highest. For the bundle sheath, DNA motif enrichment and gene coexpression analyses suggested members of the DNA binding with one finger (DOF) and MADS (MINICHROMOSOME MAINTENANCE FACTOR 1/AGAMOUS/DEFICIENS/Serum Response Factor)-domain transcription factor families mediate diurnal fluctuations in C4 gene expression, while trans-activation assays in planta confirmed their ability to activate promoter fragments from bundle sheath expressed genes. The work thus identifies transcriptional regulators and peaks in cell-specific C4 gene expression coincident with maximum rates of photosynthesis in the maize leaf at midday.
Asunto(s)
Fotosíntesis , Zea mays , Zea mays/genética , Zea mays/metabolismo , Fotosíntesis/genética , Factores de Transcripción/metabolismo , Regiones Promotoras Genéticas/genética , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Expresión GénicaRESUMEN
Plant leaves contain multiple cell types which achieve distinct characteristics whilst still coordinating development within the leaf. The bundle sheath possesses larger individual cells and lower chloroplast content than the adjacent mesophyll, but how this morphology is achieved remains unknown. To identify regulatory mechanisms determining bundle sheath cell morphology we tested the effects of perturbing environmental (light) and endogenous signals (hormones) during leaf development of Oryza sativa (rice). Total chloroplast area in bundle sheath cells was found to increase with cell size as in the mesophyll but did not maintain a 'set-point' relationship, with the longest bundle sheath cells demonstrating the lowest chloroplast content. Application of exogenous cytokinin and gibberellin significantly altered the relationship between cell size and chloroplast biosynthesis in the bundle sheath, increasing chloroplast content of the longest cells. Delayed exposure to light reduced the mean length of bundle sheath cells but increased corresponding leaf length, whereas premature light reduced final leaf length but did not affect bundle sheath cells. This suggests that the plant hormones cytokinin and gibberellin are regulators of the bundle sheath cell-chloroplast relationship and that final bundle sheath length may potentially be affected by light-mediated control of exit from the cell cycle.
Asunto(s)
Cloroplastos , Citocininas , Giberelinas , Luz , Oryza , Reguladores del Crecimiento de las Plantas , Hojas de la Planta , Oryza/crecimiento & desarrollo , Oryza/efectos de la radiación , Oryza/citología , Hojas de la Planta/crecimiento & desarrollo , Hojas de la Planta/efectos de la radiación , Citocininas/metabolismo , Citocininas/farmacología , Giberelinas/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Cloroplastos/metabolismo , Forma de la Célula/efectos de la radiación , Factores de Tiempo , Tamaño de la Célula/efectos de la radiaciónRESUMEN
When exposed to high light, plants produce reactive oxygen species (ROS). In Arabidopsis thaliana, local stress such as excess heat or light initiates a systemic ROS wave in phloem and xylem cells dependent on NADPH oxidase/respiratory burst oxidase homolog (RBOH) proteins. In the case of excess light, although the initial local accumulation of ROS preferentially takes place in bundle-sheath strands, little is known about how this response takes place. Using rice and the ROS probes diaminobenzidine and 2',7'-dichlorodihydrofluorescein diacetate, we found that, after exposure to high light, ROS were produced more rapidly in bundle-sheath strands than mesophyll cells. This response was not affected either by CO2 supply or photorespiration. Consistent with these findings, deep sequencing of messenger RNA (mRNA) isolated from mesophyll or bundle-sheath strands indicated balanced accumulation of transcripts encoding all major components of the photosynthetic apparatus. However, transcripts encoding several isoforms of the superoxide/H2O2-producing enzyme NADPH oxidase were more abundant in bundle-sheath strands than mesophyll cells. ROS production in bundle-sheath strands was decreased in mutant alleles of the bundle-sheath strand preferential isoform of OsRBOHA and increased when it was overexpressed. Despite the plethora of pathways able to generate ROS in response to excess light, NADPH oxidase-mediated accumulation of ROS in the rice bundle-sheath strand was detected in etiolated leaves lacking chlorophyll. We conclude that photosynthesis is not necessary for the local ROS response to high light but is in part mediated by NADPH oxidase activity.
Asunto(s)
Luz , NADPH Oxidasas/metabolismo , Oryza/enzimología , Oryza/efectos de la radiación , Fotosíntesis/efectos de la radiación , Haz Vascular de Plantas/enzimología , Haz Vascular de Plantas/fisiología , Especies Reactivas de Oxígeno/metabolismo , Bencidinas/metabolismo , Dióxido de Carbono/metabolismo , Regulación de la Expresión Génica de las Plantas , Oryza/genética , Oryza/fisiología , Oxígeno/metabolismo , Fotosíntesis/genética , Hojas de la Planta/genética , Hojas de la Planta/efectos de la radiación , Haz Vascular de Plantas/efectos de la radiación , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transcriptoma/genéticaRESUMEN
Reactive oxygen species (ROS) and the photoreceptor protein phytochrome B (phyB) play a key role in plant acclimation to stress. However, how phyB that primarily functions in the nuclei impacts ROS signaling mediated by respiratory burst oxidase homolog (RBOH) proteins that reside on the plasma membrane, during stress, is unknown. Arabidopsis thaliana and Oryza sativa mutants, RNA-Seq, bioinformatics, biochemistry, molecular biology, and whole-plant ROS imaging were used to address this question. Here, we reveal that phyB and RBOHs function as part of a key regulatory module that controls apoplastic ROS production, stress-response transcript expression, and plant acclimation in response to excess light stress. We further show that phyB can regulate ROS production during stress even if it is restricted to the cytosol and that phyB, respiratory burst oxidase protein D (RBOHD), and respiratory burst oxidase protein F (RBOHF) coregulate thousands of transcripts in response to light stress. Surprisingly, we found that phyB is also required for ROS accumulation in response to heat, wounding, cold, and bacterial infection. Our findings reveal that phyB plays a canonical role in plant responses to biotic and abiotic stresses, regulating apoplastic ROS production, possibly while at the cytosol, and that phyB and RBOHD/RBOHF function in the same regulatory pathway.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/metabolismo , Fitocromo B/genética , Fitocromo B/metabolismo , Oxígeno/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Arabidopsis/metabolismo , Estrés Fisiológico , Regulación de la Expresión Génica de las PlantasRESUMEN
C4 photosynthesis has evolved multiple times in the angiosperms and typically involves alterations to the biochemistry, cell biology and development of leaves. One common modification found in C4 plants compared with the ancestral C3 state is an increase in vein density such that the leaf contains a larger proportion of bundle sheath cells. Recent findings indicate that there may be significant intraspecific variation in traits such as vein density in C4 plants but to use such natural variation for trait-mapping, rapid phenotyping would be required. Here we report a high-throughput method to quantify vein density that leverages the bundle sheath-specific accumulation of starch found in C4 species. Starch staining allowed high-contrast images to be acquired permitting image analysis with MATLAB- and Python-based programmes. The method works for dicotyledons and monocotolydons. We applied this method to Gynandropsis gynandra where significant variation in vein density was detected between natural accessions, and Zea mays where no variation was apparent in the genotypically diverse lines assessed. We anticipate this approach will be useful to map genes controlling vein density in C4 species demonstrating natural variation for this trait.
Asunto(s)
Magnoliopsida , Almidón , Almidón/metabolismo , Plantas/metabolismo , Hojas de la Planta/metabolismo , Zea mays/metabolismo , Fotosíntesis/genética , Magnoliopsida/metabolismoRESUMEN
Leaves comprise multiple cell types but our knowledge of the patterns of gene expression that underpin their functional specialization is fragmentary. Our understanding and ability to undertake the rational redesign of these cells is therefore limited. We aimed to identify genes associated with the incompletely understood bundle sheath of C3 plants, which represents a key target associated with engineering traits such as C4 photosynthesis into Oryza sativa (rice). To better understand the veins, bundle sheath and mesophyll cells of rice, we used laser capture microdissection followed by deep sequencing. Gene expression of the mesophyll is conditioned to allow coenzyme metabolism and redox homeostasis, as well as photosynthesis. In contrast, the bundle sheath is specialized in water transport, sulphur assimilation and jasmonic acid biosynthesis. Despite the small chloroplast compartment of bundle sheath cells, substantial photosynthesis gene expression was detected. These patterns of gene expression were not associated with the presence or absence of specific transcription factors in each cell type, but were instead associated with gradients in expression across the leaf. Comparative analysis with C3 Arabidopsis identified a small gene set preferentially expressed in the bundle sheath cells of both species. This gene set included genes encoding transcription factors from 14 orthogroups and proteins allowing water transport, sulphate assimilation and jasmonic acid synthesis. The most parsimonious explanation for our findings is that bundle sheath cells from the last common ancestor of rice and Arabidopsis were specialized in this manner, and as the species diverged these patterns of gene expression have been maintained.
Asunto(s)
Ciclopentanos/metabolismo , Regulación de la Expresión Génica de las Plantas , Oryza/metabolismo , Oxilipinas/metabolismo , Azufre/metabolismo , Agua/metabolismo , Arabidopsis/genética , Transporte Biológico/genética , Transporte Biológico/fisiología , Células del Mesófilo/metabolismo , Nitrógeno/metabolismo , Oryza/genética , Oryza/fisiología , Fotosíntesis , Hojas de la Planta/citología , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
In biological discovery and engineering research, there is a need to spatially and/or temporally regulate transgene expression. However, the limited availability of promoter sequences that are uniquely active in specific tissue-types and/or at specific times often precludes co-expression of multiple transgenes in precisely controlled developmental contexts. Here, we developed a system for use in rice that comprises synthetic designer transcription activator-like effectors (dTALEs) and cognate synthetic TALE-activated promoters (STAPs). The system allows multiple transgenes to be expressed from different STAPs, with the spatial and temporal context determined by a single promoter that drives expression of the dTALE. We show that two different systems-dTALE1-STAP1 and dTALE2-STAP2-can activate STAP-driven reporter gene expression in stable transgenic rice lines, with transgene transcript levels dependent on both dTALE and STAP sequence identities. The relative strength of individual STAP sequences is consistent between dTALE1 and dTALE2 systems but differs between cell-types, requiring empirical evaluation in each case. dTALE expression leads to off-target activation of endogenous genes but the number of genes affected is substantially less than the number impacted by the somaclonal variation that occurs during the regeneration of transformed plants. With the potential to design fully orthogonal dTALEs for any genome of interest, the dTALE-STAP system thus provides a powerful approach to fine-tune the expression of multiple transgenes, and to simultaneously introduce different synthetic circuits into distinct developmental contexts.
Asunto(s)
Oryza , Genes Reporteros , Oryza/genética , Plantas/genética , Plantas Modificadas Genéticamente/genética , Regiones Promotoras Genéticas/genética , Transgenes/genéticaRESUMEN
Chloroplasts are best known for their role in photosynthesis, but they also allow nitrogen and sulphur assimilation, amino acid, fatty acid, nucleotide and hormone synthesis. How chloroplasts develop is therefore relevant to these diverse and fundamental biological processes, but also to attempts at their rational redesign. Light is strictly required for chloroplast formation in all angiosperms and directly regulates the expression of hundreds of chloroplast-related genes. Light also modulates the levels of several hormones including brassinosteriods, cytokinins, auxins and gibberellins, which themselves control chloroplast development particularly during early stages of plant development. Transcription factors such as GOLDENLIKE1&2 (GLK1&2), GATA NITRATE-INDUCIBLE CARBON METABOLISM-INVOLVED (GNC) and CYTOKININ-RESPONSIVE GATA FACTOR 1 (CGA1) act downstream of both light and phytohormone signalling to regulate chloroplast development. Thus, in green tissues transcription factors, light signalling and hormone signalling form a complex network regulating the transcription of chloroplast- and photosynthesis-related genes to control the development and number of chloroplasts per cell. We use this conceptual framework to identify points of regulation that could be harnessed to modulate chloroplast abundance and increase photosynthetic efficiency of crops, and to highlight future avenues to overcome gaps in current knowledge.
Asunto(s)
Proteínas de Arabidopsis , Viridiplantae , Proteínas de Arabidopsis/metabolismo , Cloroplastos/metabolismo , Regulación de la Expresión Génica de las Plantas , Hormonas/metabolismo , Luz , Fotosíntesis/genética , Hojas de la Planta/fisiología , Viridiplantae/metabolismoRESUMEN
Reducing photorespiration in C3 crops could significantly increase rates of photosynthesis and yield. One method to achieve this would be to integrate C4 photosynthesis into C3 species. This objective is challenging as it involves engineering incompletely understood traits into C3 leaves, including complex changes to their biochemistry, cell biology, and anatomy. Quantitative genetics and selective breeding offer underexplored routes to identify regulators of these processes. We first review examples of natural intraspecific variation in C4 photosynthesis as well as the potential for hybridization between C3 and C4 species. We then discuss how quantitative genetic approaches including artificial selection and genome-wide association could be used to better understand the C4 syndrome and in so doing guide the engineering of the C4 pathway into C3 crops.
Asunto(s)
Estudio de Asociación del Genoma Completo , Fitomejoramiento , Productos Agrícolas/genética , Fotosíntesis/genética , Hojas de la Planta/metabolismoRESUMEN
The majority of plants use C3 photosynthesis, but over 60 independent lineages of angiosperms have evolved the C4 pathway. In most C4 species, photosynthesis gene expression is compartmented between mesophyll and bundle-sheath cells. We performed DNaseI sequencing to identify genome-wide profiles of transcription factor binding in leaves of the C4 grasses Zea mays, Sorghum bicolor, and Setaria italica as well as C3 Brachypodium distachyon In C4 species, while bundle-sheath strands and whole leaves shared similarity in the broad regions of DNA accessible to transcription factors, the short sequences bound varied. Transcription factor binding was prevalent in gene bodies as well as promoters, and many of these sites could represent duons that influence gene regulation in addition to amino acid sequence. Although globally there was little correlation between any individual DNaseI footprint and cell-specific gene expression, within individual species transcription factor binding to the same motifs in multiple genes provided evidence for shared mechanisms governing C4 photosynthesis gene expression. Furthermore, interspecific comparisons identified a small number of highly conserved transcription factor binding sites associated with leaves from species that diverged around 60 million years ago. These data therefore provide insight into the architecture associated with C4 photosynthesis gene expression in particular and characteristics of transcription factor binding in cereal crops in general.
Asunto(s)
Fotosíntesis/genética , Proteínas de Plantas/metabolismo , Poaceae/genética , Factores de Transcripción/metabolismo , Brachypodium/genética , Desoxirribonucleasa I , Eucromatina/genética , Eucromatina/metabolismo , Evolución Molecular , Regulación de la Expresión Génica de las Plantas/genética , Genoma de Planta , Motivos de Nucleótidos , Fotosíntesis/fisiología , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Filogenia , Hojas de la Planta/enzimología , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Regiones Promotoras Genéticas , Análisis de Secuencia de ADN , Setaria (Planta)/genética , Setaria (Planta)/metabolismo , Sorghum/genética , Sorghum/metabolismo , Zea mays/genética , Zea mays/metabolismoRESUMEN
The engineering of C4 photosynthetic activity into the C3 plant rice has the potential to nearly double rice yields. To engineer a two-cell photosynthetic system in rice, the rice bundle sheath (BS) must be rewired to enhance photosynthetic capacity. Here, we show that BS chloroplast biogenesis is enhanced when the transcriptional activator, Oryza sativa Cytokinin GATA transcription factor 1 (OsCGA1), is driven by a vascular specific promoter. Ectopic expression of OsCGA1 resulted in increased BS chloroplast planar area and increased expression of photosynthesis-associated nuclear genes (PhANG), required for the biogenesis of photosynthetically active chloroplasts in BS cells of rice. A further refinement using a DNAse dead Cas9 (dCas9) activation module driven by the same cell-type specific promoter, directed enhanced chloroplast development of the BS cells when gRNA sequences were delivered by the dCas9 module to the promoter of the endogenous OsCGA1 gene. Single gRNA expression was sufficient to mediate the transactivation of both the endogenous gene and a transgenic GUS reporter fused with OsCGA1 promoter. Our results illustrate the potential for tissue-specific dCas9-activation and the co-regulation of genes needed for multistep engineering of C4 rice.
Asunto(s)
Oryza , Cloroplastos/genética , Cloroplastos/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Oryza/genética , Fotosíntesis/genética , Hojas de la Planta , Regiones Promotoras Genéticas/genéticaRESUMEN
If the highly efficient C4 photosynthesis pathway could be transferred to crops with the C3 pathway there could be yield gains of up to 50%. It has been proposed that the multiple metabolic and developmental modifications associated with C4 photosynthesis are underpinned by relatively few master regulators that have allowed the evolution of C4 photosynthesis more than 60 times in flowering plants. Here we identify a component of one such regulator that consists of a pair of cis-elements located in coding sequence of multiple genes that are preferentially expressed in bundle sheath cells of C4 leaves. These motifs represent duons as they play a dual role in coding for amino acids as well as controlling the spatial patterning of gene expression associated with the C4 leaf. They act to repress transcription of C4 photosynthesis genes in mesophyll cells. These duons are also present in the C3 model Arabidopsis thaliana, and, in fact, are conserved in all land plants and even some algae that use C3 photosynthesis. C4 photosynthesis therefore appears to have coopted an ancient regulatory code to generate the spatial patterning of gene expression that is a hallmark of C4 photosynthesis. This intragenic transcriptional regulatory sequence could be exploited in the engineering of efficient photosynthesis of crops.