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1.
Plant Commun ; : 100984, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38845198

RESUMO

The soybean root system is complex. In addition to being composed of various cell types, the soybean root system includes the primary root, the lateral roots, and the nodule, an organ in which mutualistic symbiosis with the N-fixing rhizobia occurs. A mature soybean root nodule is characterized by a central infection zone where the atmospheric nitrogen is fixed and assimilated by the symbiont, resulting from the close cooperation between the plant cell and the bacteria. To date, the transcriptome of individual cells isolated from developing soybean nodules has been established, but the transcriptomic signatures of the cells of the mature soybean nodule have not yet been characterized. Applying single nucleus RNA-seq and Molecular CartographyTM technologies, we precisely characterized the transcriptomic signature of the soybean root and mature nodule cell types and revealed the co-existence of different sub-populations of B. diazoefficiens-infected cells in the mature soybean nodule including those actively involved in nitrogen fixation, and those engaged in senescence. The mining of the single cell-resolution nodule transcriptome atlas and associated gene co-expression network confirmed the role of known nodulation-related genes and identified new genes controlling the nodulation process. For instance, we functionally characterized the role of GmFWL3, a plasma membrane microdomain-associated protein controlling rhizobia infection. Our study reveals the unique cellular complexity of the mature soybean nodule and helps redefine the concept of cell types when considering the infection zone of the soybean nodule.

2.
bioRxiv ; 2024 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-38352530

RESUMO

Screening a transposon-mutagenized soybean population led to the discovery of a recessively inherited chlorotic phenotype. This "vir1" phenotype results in smaller stature, weaker stems, and a smaller root system with smaller nodules. Genome sequencing identified 15 candidate genes with mutations likely to result in a loss of function. Amplicon sequencing of a segregating population was then used to narrow the list to a single candidate mutation, a single-base change in Glyma.07G102300 that disrupts splicing of the second intron. Single cell transcriptomic profiling indicates that this gene is expressed primarily in mesophyll cells and RNA sequencing data indicates it is upregulated in germinating seedlings by cold stress. Previous studies have shown that mutations to Os05g34040, the rice homolog of Glyma.07G102300, produced a chlorotic phenotype that was more pronounced in cool temperatures. Growing soybean vir1 mutants at lower temperatures also resulted in a more severe phenotype. In addition, transgenic expression of wild type Glyma.07G102300 in the knockout mutant of the Arabidopsis homolog At4930720 rescues the chlorotic phenotype, further supporting the hypothesis that the mutation in Glyma.07G102300 is causal of the vir1 phenotype.

4.
Mol Plant ; 15(12): 1868-1888, 2022 12 05.
Artigo em Inglês | MEDLINE | ID: mdl-36321199

RESUMO

Medicago truncatula is a model legume species that has been studied for decades to understand the symbiotic relationship between legumes and soil bacteria collectively named rhizobia. This symbiosis called nodulation is initiated in roots with the infection of root hair cells by the bacteria, as well as the initiation of nodule primordia from root cortical, endodermal, and pericycle cells, leading to the development of a new root organ, the nodule, where bacteria fix and assimilate the atmospheric dinitrogen for the benefit of the plant. Here, we report the isolation and use of the nuclei from mock and rhizobia-inoculated roots for the single nuclei RNA-seq (sNucRNA-seq) profiling to gain a deeper understanding of early responses to rhizobial infection in Medicago roots. A gene expression map of the Medicago root was generated, comprising 25 clusters, which were annotated as specific cell types using 119 Medicago marker genes and orthologs to Arabidopsis cell-type marker genes. A focus on root hair, cortex, endodermis, and pericycle cell types, showing the strongest differential regulation in response to a short-term (48 h) rhizobium inoculation, revealed not only known genes and functional pathways, validating the sNucRNA-seq approach, but also numerous novel genes and pathways, allowing a comprehensive analysis of early root symbiotic responses at a cell type-specific level.


Assuntos
Medicago truncatula , Medicago truncatula/genética
5.
Plant Sci ; 325: 111486, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36202294

RESUMO

Plant single-cell RNA-seq technology quantifies the abundance of plant transcripts at a single-cell resolution. Deciphering the transcriptomes of each plant cell, their regulation during plant cell development, and their response to environmental stresses will support the functional study of genes, the establishment of precise transcriptional programs, the prediction of more accurate gene regulatory networks, and, in the long term, the design of de novo gene pathways to enhance selected crop traits. In this review, we will discuss the opportunities, challenges, and problems, and share tentative solutions associated with the generation and analysis of plant single-cell transcriptomes. We will discuss the benefit and limitations of using plant protoplasts vs. nuclei to conduct single-cell RNA-seq experiments on various plant species and organs, the functional annotation of plant cell types based on their transcriptomic profile, the characterization of the dynamic regulation of the plant genes during cell development or in response to environmental stress, the need to characterize and integrate additional layers of -omics datasets to capture new molecular modalities at the single-cell level and reveal their causalities, the deposition and access to single-cell datasets, and the accessibility of this technology to plant scientists.


Assuntos
Plantas , Transcriptoma , RNA-Seq , Análise de Sequência de RNA , Plantas/genética , Tecnologia , Biologia , Perfilação da Expressão Gênica
6.
Membranes (Basel) ; 12(9)2022 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-36135893

RESUMO

Membrane proteins work in large complexes to perceive and transduce external signals and to trigger a cellular response leading to the adaptation of the cells to their environment. Biochemical assays have been extensively used to reveal the interaction between membrane proteins. However, such analyses do not reveal the unique and complex composition of the membrane proteins of the different plant cell types. Here, we conducted a comprehensive analysis of the expression of Arabidopsis membrane proteins in the different cell types composing the root. Specifically, we analyzed the expression of genes encoding membrane proteins interacting in large complexes. We found that the transcriptional profiles of membrane protein-encoding genes differ between Arabidopsis root cell types. This result suggests that different cell types are characterized by specific sets of plasma membrane proteins, which are likely a reflection of their unique biological functions and interactions. To further explore the complexity of the Arabidopsis root cell membrane proteomes, we conducted a co-expression analysis of genes encoding interacting membrane proteins. This study confirmed previously reported interactions between membrane proteins, suggesting that the co-expression of genes at the single cell-type level can be used to support protein network predictions.

7.
Sci Rep ; 11(1): 15725, 2021 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-34344949

RESUMO

The most studied DNA methylation pathway in plants is the RNA Directed DNA Methylation (RdDM), a conserved mechanism that involves the role of noncoding RNAs to control the expansion of the noncoding genome. Genome-wide DNA methylation levels have been reported to correlate with genome size. However, little is known about the catalog of noncoding RNAs and the impact on DNA methylation in small plant genomes with reduced noncoding regions. Because of the small length of intergenic regions in the compact genome of the carnivorous plant Utricularia gibba, we investigated its repertoire of noncoding RNA and DNA methylation landscape. Here, we report that, compared to other angiosperms, U. gibba has an unusual distribution of small RNAs and reduced global DNA methylation levels. DNA methylation was determined using a novel strategy based on long-read DNA sequencing with the Pacific Bioscience platform and confirmed by whole-genome bisulfite sequencing. Moreover, some key genes involved in the RdDM pathway may not represented by compensatory paralogs or comprise truncated proteins, for example, U. gibba DICER-LIKE 3 (DCL3), encoding a DICER endonuclease that produces 24-nt small-interfering RNAs, has lost key domains required for complete function. Our results unveil that a truncated DCL3 correlates with a decreased proportion of 24-nt small-interfering RNAs, low DNA methylation levels, and developmental abnormalities during female gametogenesis in U. gibba. Alterations in female gametogenesis are reminiscent of RdDM mutant phenotypes in Arabidopsis thaliana. It would be interesting to further study the biological implications of the DCL3 truncation in U. gibba, as it could represent an initial step in the evolution of RdDM pathway in compact genomes.


Assuntos
Metilação de DNA , Endonucleases/genética , Endonucleases/metabolismo , Gametogênese , Lamiales/fisiologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Mutação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , RNA de Plantas , RNA não Traduzido/metabolismo , Ribonuclease III/genética , Ribonuclease III/metabolismo
8.
Int J Mol Sci ; 21(21)2020 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-33171770

RESUMO

Phosphate (Pi) is a pivotal nutrient that constraints plant development and productivity in natural ecosystems. Land colonization by plants, more than 470 million years ago, evolved adaptive mechanisms to conquer Pi-scarce environments. However, little is known about the molecular basis underlying such adaptations at early branches of plant phylogeny. To shed light on how early divergent plants respond to Pi limitation, we analyzed the morpho-physiological and transcriptional dynamics of Marchantia polymorpha upon Pi starvation. Our phylogenomic analysis highlights some gene networks present since the Chlorophytes and others established in the Streptophytes (e.g., PHR1-SPX1 and STOP1-ALMT1, respectively). At the morpho-physiological level, the response is characterized by the induction of phosphatase activity, media acidification, accumulation of auronidins, reduction of internal Pi concentration, and developmental modifications of rhizoids. The transcriptional response involves the induction of MpPHR1, Pi transporters, lipid turnover enzymes, and MpMYB14, which is an essential transcription factor for auronidins biosynthesis. MpSTOP2 up-regulation correlates with expression changes in genes related to organic acid biosynthesis and transport, suggesting a preference for citrate exudation. An analysis of MpPHR1 binding sequences (P1BS) shows an enrichment of this cis regulatory element in differentially expressed genes. Our study unravels the strategies, at diverse levels of organization, exerted by M. polymorpha to cope with low Pi availability.


Assuntos
Marchantia/genética , Marchantia/metabolismo , Fosfatos/metabolismo , Ecossistema , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/genética , Redes Reguladoras de Genes/efeitos dos fármacos , Redes Reguladoras de Genes/genética , Hepatófitas/metabolismo , Filogenia , Fatores de Transcrição/metabolismo
9.
Dev Biol ; 442(1): 28-39, 2018 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-29705332

RESUMO

In plants, the best characterized plant regeneration process is de novo organogenesis. This type of regeneration is characterized by the formation of a multicellular structure called callus. Calli are induced via phytohormone treatment of plant sections. The callus formation in plants like Agave species with Crassulacean Acid Metabolism (CAM) is poorly studied. In this study, we induced callus formation from Agave salmiana leaves and describe cell arrangement in this tissue. Moreover, we determined and analyzed the transcriptional program of calli, as well as those of differentiated root and leaf tissues, by using RNA-seq. We were able to reconstruct 170,844 transcripts of which 40,644 have a full Open Reading Frame (ORF). The global profile obtained by Next Generation Sequencing (NGS) reveals that several callus-enriched protein coding transcripts are orthologs of previously reported factors highly expressed in Arabidopsis calli. At least 62 genes were differentially expressed in Agave calli, 50 of which were up-regulated. Several of these are actively involved in the perception of, and response to, auxin and cytokinin. Not only are these the first results for the A. salmiana callus, but they provide novel data from roots and leaves of this Agave species, one of the largest non-tree plants in nature.


Assuntos
Agave/genética , Organogênese Vegetal/genética , Regeneração/genética , Crassulaceae/genética , Citocininas/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas/genética , Sequenciamento de Nucleotídeos em Larga Escala , Ácidos Indolacéticos/metabolismo , Organogênese Vegetal/fisiologia , Reguladores de Crescimento de Plantas/genética , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Transcriptoma/genética
10.
Proc Natl Acad Sci U S A ; 114(22): E4435-E4441, 2017 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-28507139

RESUMO

Utricularia gibba, the humped bladderwort, is a carnivorous plant that retains a tiny nuclear genome despite at least two rounds of whole genome duplication (WGD) since common ancestry with grapevine and other species. We used a third-generation genome assembly with several complete chromosomes to reconstruct the two most recent lineage-specific ancestral genomes that led to the modern U. gibba genome structure. Patterns of subgenome dominance in the most recent WGD, both architectural and transcriptional, are suggestive of allopolyploidization, which may have generated genomic novelty and led to instantaneous speciation. Syntenic duplicates retained in polyploid blocks are enriched for transcription factor functions, whereas gene copies derived from ongoing tandem duplication events are enriched in metabolic functions potentially important for a carnivorous plant. Among these are tandem arrays of cysteine protease genes with trap-specific expression that evolved within a protein family known to be useful in the digestion of animal prey. Further enriched functions among tandem duplicates (also with trap-enhanced expression) include peptide transport (intercellular movement of broken-down prey proteins), ATPase activities (bladder-trap acidification and transmembrane nutrient transport), hydrolase and chitinase activities (breakdown of prey polysaccharides), and cell-wall dynamic components possibly associated with active bladder movements. Whereas independently polyploid Arabidopsis syntenic gene duplicates are similarly enriched for transcriptional regulatory activities, Arabidopsis tandems are distinct from those of U. gibba, while still metabolic and likely reflecting unique adaptations of that species. Taken together, these findings highlight the special importance of tandem duplications in the adaptive landscapes of a carnivorous plant genome.


Assuntos
Carnivoridade/fisiologia , Genoma de Planta , Lamiales/genética , Lamiales/fisiologia , Adaptação Fisiológica/genética , Cisteína Proteases/química , Cisteína Proteases/genética , Evolução Molecular , Duplicação Gênica , Modelos Moleculares , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Poliploidia , Análise de Sequência de DNA , Sintenia
11.
Plant Signal Behav ; 11(5): e1173300, 2016 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-27185363

RESUMO

Phosphate (Pi) limitation is a constraint for plant growth in many natural and agricultural ecosystems. Plants possess adaptive mechanisms that enable them to cope with conditions of limited Pi supply, including a highly regulated genetic program controlling the expression of genes involved in different metabolic, signaling and development processes of plants. Recently, we showed that in response to phosphate limitation Arabidopsis thaliana sets specific DNA methylation patterns of genic features that often correlated with changes in gene expression. Our findings included, dynamic methylation changes in response to phosphate starvation and the observation that the expression of genes encoding DNA methyltransferases appear to be directly controlled by the key regulator PHOSPHATE RESPONSE 1 (PHR1). These results provide insight into how epigenetic marks can influence plant genomes and transcriptional programs to respond and adapt to harsh conditions. Here we present an analysis of DNA methylation in the upstream regions of low Pi responsive genes in Arabidopsis seedlings exposed to low Pi conditions. We found that hypo- and hyper-methylation in the vicinity of cognate binding sites for transcription factors known to regulate the phosphate starvation response clearly correlates with increased or decreased expression of low-Pi responsive genes.


Assuntos
Arabidopsis/genética , Metilação de DNA/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Fosfatos/deficiência , Sequências Reguladoras de Ácido Nucleico/genética , Arabidopsis/efeitos dos fármacos , Sequência de Bases , Metilação de DNA/efeitos dos fármacos , Perfilação da Expressão Gênica , Ontologia Genética , Motivos de Nucleotídeos/genética , Fosfatos/farmacologia
12.
Proc Natl Acad Sci U S A ; 112(52): E7293-302, 2015 Dec 29.
Artigo em Inglês | MEDLINE | ID: mdl-26668375

RESUMO

Phosphate (Pi) availability is a significant limiting factor for plant growth and productivity in both natural and agricultural systems. To cope with such limiting conditions, plants have evolved a myriad of developmental and biochemical strategies to enhance the efficiency of Pi acquisition and assimilation to avoid nutrient starvation. In the past decade, these responses have been studied in detail at the level of gene expression; however, the possible epigenetic components modulating plant Pi starvation responses have not been thoroughly investigated. Here, we report that an extensive remodeling of global DNA methylation occurs in Arabidopsis plants exposed to low Pi availability, and in many instances, this effect is related to changes in gene expression. Modifications in methylation patterns within genic regions were often associated with transcriptional activation or repression, revealing the important role of dynamic methylation changes in modulating the expression of genes in response to Pi starvation. Moreover, Arabidopsis mutants affected in DNA methylation showed that changes in DNA methylation patterns are required for the accurate regulation of a number of Pi-starvation-responsive genes and that DNA methylation is necessary to establish proper morphological and physiological phosphate starvation responses.


Assuntos
Arabidopsis/genética , Metilação de DNA , Epigênese Genética , Epigenômica/métodos , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Fosfatos/metabolismo , Adaptação Fisiológica/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , DNA (Citosina-5-)-Metiltransferases/genética , Mutação , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase Via Transcriptase Reversa
13.
Nature ; 498(7452): 94-8, 2013 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-23665961

RESUMO

It has been argued that the evolution of plant genome size is principally unidirectional and increasing owing to the varied action of whole-genome duplications (WGDs) and mobile element proliferation. However, extreme genome size reductions have been reported in the angiosperm family tree. Here we report the sequence of the 82-megabase genome of the carnivorous bladderwort plant Utricularia gibba. Despite its tiny size, the U. gibba genome accommodates a typical number of genes for a plant, with the main difference from other plant genomes arising from a drastic reduction in non-genic DNA. Unexpectedly, we identified at least three rounds of WGD in U. gibba since common ancestry with tomato (Solanum) and grape (Vitis). The compressed architecture of the U. gibba genome indicates that a small fraction of intergenic DNA, with few or no active retrotransposons, is sufficient to regulate and integrate all the processes required for the development and reproduction of a complex organism.


Assuntos
Evolução Molecular , Genoma de Planta/genética , Magnoliopsida/genética , DNA Intergênico/genética , Duplicação Gênica/genética , Genes de Plantas/genética , Modelos Genéticos , Solanum/genética , Sintenia/genética , Vitis/genética
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