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1.
FASEB J ; 38(1): e23339, 2024 01.
Artigo em Inglês | MEDLINE | ID: mdl-38069905

RESUMO

Being overweight exacerbates various metabolic diseases, necessitating the identification of target molecules for obesity control. In the current study, we investigated common physiological features related to metabolism in mice with low weight gain: (1) G protein-coupled receptor, family C, group 5, member B-knockout; (2) gastric inhibitory polypeptide receptor-knockout; and (3) Iroquois-related homeobox 3-knockout. Moreover, we explored genes involved in metabolism by analyzing differentially expressed genes (DEGs) between low-weight gain mice and the respective wild-type control mice. The common characteristics of the low-weight gain mice were low inguinal white adipose tissue (iWAT) and liver weight despite similar food intake along with lower blood leptin levels and high energy expenditure. The DEGs of iWAT, epididymal (gonadal) WAT, brown adipose tissue, muscle, liver, hypothalamus, and hippocampus common to these low-weight gain mice were designated as candidate genes associated with metabolism. One such gene tetraspanin 7 (Tspan7) from the iWAT was validated using knockout and overexpressing mouse models. Mice with low Tspan7 expression gained more weight, while those with high Tspan7 expression gained less weight, confirming the involvement of the Tspan7 gene in weight regulation. Collectively, these findings suggest that the candidate gene list generated in this study contains potential target molecules for obesity regulation. Further validation and additional data from low-weight gain mice will aid in understanding the molecular mechanisms associated with obesity.


Assuntos
Tecido Adiposo Marrom , Obesidade , Camundongos , Animais , Obesidade/genética , Obesidade/metabolismo , Tecido Adiposo Marrom/metabolismo , Aumento de Peso/genética , Tecido Adiposo Branco/metabolismo , Metabolismo Energético/genética , Fenótipo , Camundongos Endogâmicos C57BL , Dieta Hiperlipídica , Camundongos Knockout
2.
Nat Commun ; 14(1): 6133, 2023 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-37783685

RESUMO

Atopic dermatitis (AD) is a skin disease that is heterogeneous both in terms of clinical manifestations and molecular profiles. It is increasingly recognized that AD is a systemic rather than a local disease and should be assessed in the context of whole-body pathophysiology. Here we show, via integrated RNA-sequencing of skin tissue and peripheral blood mononuclear cell (PBMC) samples along with clinical data from 115 AD patients and 14 matched healthy controls, that specific clinical presentations associate with matching differential molecular signatures. We establish a regression model based on transcriptome modules identified in weighted gene co-expression network analysis to extract molecular features associated with detailed clinical phenotypes of AD. The two main, qualitatively differential skin manifestations of AD, erythema and papulation are distinguished by differential immunological signatures. We further apply the regression model to a longitudinal dataset of 30 AD patients for personalized monitoring, highlighting patient heterogeneity in disease trajectories. The longitudinal features of blood tests and PBMC transcriptome modules identify three patient clusters which are aligned with clinical severity and reflect treatment history. Our approach thus serves as a framework for effective clinical investigation to gain a holistic view on the pathophysiology of complex human diseases.


Assuntos
Dermatite Atópica , Humanos , Dermatite Atópica/genética , Transcriptoma , Leucócitos Mononucleares , Pele , Fenótipo
3.
Nature ; 621(7978): 389-395, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37648852

RESUMO

Insulin resistance is the primary pathophysiology underlying metabolic syndrome and type 2 diabetes1,2. Previous metagenomic studies have described the characteristics of gut microbiota and their roles in metabolizing major nutrients in insulin resistance3-9. In particular, carbohydrate metabolism of commensals has been proposed to contribute up to 10% of the host's overall energy extraction10, thereby playing a role in the pathogenesis of obesity and prediabetes3,4,6. Nevertheless, the underlying mechanism remains unclear. Here we investigate this relationship using a comprehensive multi-omics strategy in humans. We combine unbiased faecal metabolomics with metagenomics, host metabolomics and transcriptomics data to profile the involvement of the microbiome in insulin resistance. These data reveal that faecal carbohydrates, particularly host-accessible monosaccharides, are increased in individuals with insulin resistance and are associated with microbial carbohydrate metabolisms and host inflammatory cytokines. We identify gut bacteria associated with insulin resistance and insulin sensitivity that show a distinct pattern of carbohydrate metabolism, and demonstrate that insulin-sensitivity-associated bacteria ameliorate host phenotypes of insulin resistance in a mouse model. Our study, which provides a comprehensive view of the host-microorganism relationships in insulin resistance, reveals the impact of carbohydrate metabolism by microbiota, suggesting a potential therapeutic target for ameliorating insulin resistance.


Assuntos
Metabolismo dos Carboidratos , Microbioma Gastrointestinal , Resistência à Insulina , Animais , Humanos , Camundongos , Diabetes Mellitus Tipo 2/metabolismo , Microbioma Gastrointestinal/fisiologia , Resistência à Insulina/fisiologia , Monossacarídeos/metabolismo , Insulina/metabolismo , Síndrome Metabólica/metabolismo , Fezes/química , Fezes/microbiologia , Metabolômica
4.
Plant Physiol ; 187(1): 158-173, 2021 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-34618135

RESUMO

Because of limited free diffusion in the cytoplasm, viruses must use active transport mechanisms to move intracellularly. Nevertheless, how the plant single-stranded DNA begomoviruses hijack the host intracytoplasmic transport machinery to move from the nucleus to the plasmodesmata remains enigmatic. Here, we identified nuclear shuttle protein (NSP)-interacting proteins from Arabidopsis (Arabidopsis thaliana) by probing a protein microarray and demonstrated that the cabbage leaf curl virus NSP, a facilitator of the nucleocytoplasmic trafficking of viral (v)DNA, interacts in planta with an endosomal vesicle-localized, plant-specific syntaxin-6 protein, designated NSP-interacting syntaxin domain-containing protein (NISP). NISP displays a proviral function, unlike the syntaxin-6 paralog AT2G18860 that failed to interact with NSP. Consistent with these findings, nisp-1 mutant plants were less susceptible to begomovirus infection, a phenotype reversed by NISP complementation. NISP-overexpressing lines accumulated higher levels of vDNA than wild-type. Furthermore, NISP interacted with an NSP-interacting GTPase (NIG) involved in NSP-vDNA nucleocytoplasmic translocation. The NISP-NIG interaction was enhanced by NSP. We also showed that endosomal NISP associates with vDNA. NISP may function as a docking site for recruiting NIG and NSP into endosomes, providing a mechanism for the intracytoplasmic translocation of the NSP-vDNA complex toward and from the cell periphery.


Assuntos
Arabidopsis , Begomovirus , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/virologia , Begomovirus/fisiologia , Núcleo Celular/metabolismo
5.
Bio Protoc ; 10(18): e3759, 2020 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-33659418

RESUMO

Highly sensitive quantitative protein profiling can play a key role in the early diagnosis of diseases, such as autoimmune diseases and cancer. We developed a modified protein-oligonucleotide conjugation method termed HaloTag-mediated barcoding, for quantifying protein molecules at a higher sensitivity than conventional protein quantification methods. This novel and efficient conjugation method can be used to prepare HaloTag-barcoded proteins using a click chemistry-based labeling technique. Here, we describe the preparation of protein-DNA complexes and detection of protein-protein interactions which can be used in a HaloTag protein barcode assay to detect an antibody. The protocol includes procedures for preparing the ligand-oligonucleotide complex, plasmid DNA preparation for protein expression, and preparation of the protein-oligonucleotide complex. The described click reaction-based protocols simplify the conventional amine-ester reaction methods which require additional steps for chromatography purification.

6.
Nucleic Acids Res ; 48(2): e8, 2020 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-31752022

RESUMO

Highly sensitive protein quantification enables the detection of a small number of protein molecules that serve as markers/triggers for various biological phenomena, such as cancer. Here, we describe the development of a highly sensitive protein quantification system called HaloTag protein barcoding. The method involves covalent linking of a target protein to a unique molecule counting oligonucleotide at a 1:1 conjugation ratio based on an azido-cycloalkyne click reaction. The sensitivity of the HaloTag-based barcoding was remarkably higher than that of a conventional luciferase assay. The HaloTag system was successfully validated by analyzing a set of protein-protein interactions, with the identification rate of 44% protein interactions between positive reference pairs reported in the literature. Desmoglein 3, the target antigen of pemphigus vulgaris, an IgG-mediated autoimmune blistering disease, was used in a HaloTag protein barcode assay to detect the anti-DSG3 antibody. The dynamic range of the assay was over 104-times wider than that of a conventional enzyme-linked immunosorbent assay (ELISA). The technology was used to detect anti-DSG3 antibody in patient samples with much higher sensitivity compared to conventional ELISA. Our detection system, with its superior sensitivity, enables earlier detection of diseases possibly allowing the initiation of care/treatment at an early disease stage.


Assuntos
Anticorpos Anti-Idiotípicos/isolamento & purificação , Desmogleína 3/isolamento & purificação , Domínios e Motivos de Interação entre Proteínas/genética , Proteínas/isolamento & purificação , Anticorpos Anti-Idiotípicos/genética , Anticorpos Anti-Idiotípicos/imunologia , Doenças Autoimunes/diagnóstico , Doenças Autoimunes/imunologia , Química Click , Cicloparafinas/química , Desmogleína 3/genética , Desmogleína 3/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Imunoglobulina G/genética , Imunoglobulina G/imunologia , Luciferases/química , Oligonucleotídeos , Proteínas/genética , Proteínas/imunologia
7.
Curr Protoc Plant Biol ; 3(3): e20071, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30106517

RESUMO

Physical interactions between proteins and other molecules can be evaluated at a proteome scale using protein arrays, a type of high-throughput pulldown assay. We developed a modified in situ protein array known as the nucleic acid programmable protein assay (NAPPA) that allows the screening of thousands of open reading frames (ORFs) at a lower cost, with less labor, and in less time than conventional protein arrays. The HaloTag-NAPPA protein array can efficiently capture proteins expressed in situ on a glass slide using the Halo high-affinity capture tag. Here, we describe the fabrication of the array using publicly available resources and detection of protein-protein interactions (PPIs) that can be used to generate a protein interactome map. The Basic Protocol includes procedures for preparing the plasmid DNA spotted on glass slides, in situ protein expression, and PPI detection. The supporting protocols outline the construction of vectors and preparation of ORF clones. © 2018 by John Wiley & Sons, Inc.


Assuntos
Análise Serial de Proteínas/métodos , Proteoma , Vetores Genéticos , Fases de Leitura Aberta
8.
Proc Natl Acad Sci U S A ; 113(29): E4238-47, 2016 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-27357687

RESUMO

Protein microarrays enable investigation of diverse biochemical properties for thousands of proteins in a single experiment, an unparalleled capacity. Using a high-density system called HaloTag nucleic acid programmable protein array (HaloTag-NAPPA), we created high-density protein arrays comprising 12,000 Arabidopsis ORFs. We used these arrays to query protein-protein interactions for a set of 38 transcription factors and transcriptional regulators (TFs) that function in diverse plant hormone regulatory pathways. The resulting transcription factor interactome network, TF-NAPPA, contains thousands of novel interactions. Validation in a benchmarked in vitro pull-down assay revealed that a random subset of TF-NAPPA validated at the same rate of 64% as a positive reference set of literature-curated interactions. Moreover, using a bimolecular fluorescence complementation (BiFC) assay, we confirmed in planta several interactions of biological interest and determined the interaction localizations for seven pairs. The application of HaloTag-NAPPA technology to plant hormone signaling pathways allowed the identification of many novel transcription factor-protein interactions and led to the development of a proteome-wide plant hormone TF interactome network.


Assuntos
Proteínas de Arabidopsis/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Fatores de Transcrição/metabolismo , Arabidopsis/metabolismo , Análise Serial de Proteínas , Mapeamento de Interação de Proteínas
9.
Sci Rep ; 6: 28941, 2016 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-27357749

RESUMO

Abscisic acid (ABA) is a plant hormone that mediates abiotic stress tolerance and regulates growth and development. ABA binds to members of the PYL/RCAR ABA receptor family that initiate signal transduction inhibiting type 2C protein phosphatases. Although crosstalk between ABA and the hormone Jasmonic Acid (JA) has been shown, the molecular entities that mediate this interaction have yet to be fully elucidated. We report a link between ABA and JA signaling through a direct interaction of the ABA receptor PYL6 (RCAR9) with the basic helix-loop-helix transcription factor MYC2. PYL6 and MYC2 interact in yeast two hybrid assays and the interaction is enhanced in the presence of ABA. PYL6 and MYC2 interact in planta based on bimolecular fluorescence complementation and co-immunoprecipitation of the proteins. Furthermore, PYL6 was able to modify transcription driven by MYC2 using JAZ6 and JAZ8 DNA promoter elements in yeast one hybrid assays. Finally, pyl6 T-DNA mutant plants show an increased sensitivity to the addition of JA along with ABA in cotyledon expansion experiments. Overall, the present study identifies a direct mechanism for transcriptional modulation mediated by an ABA receptor different from the core ABA signaling pathway, and a putative mechanistic link connecting ABA and JA signaling pathways.


Assuntos
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Ácidos Linoleicos/metabolismo , Transdução de Sinais , Arabidopsis/genética , Peptídeos e Proteínas de Sinalização Intracelular , Ligação Proteica , Mapeamento de Interação de Proteínas , Nicotiana/genética , Técnicas do Sistema de Duplo-Híbrido
10.
Plant Cell ; 22(10): 3280-94, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20889913

RESUMO

Rice (Oryza sativa) endosperm accumulates a massive amount of storage starch and storage proteins during seed development. However, little is known about the regulatory system involved in the production of storage substances. The rice flo2 mutation resulted in reduced grain size and starch quality. Map-based cloning identified FLOURY ENDOSPERM2 (FLO2), a member of a novel gene family conserved in plants, as the gene responsible for the rice flo2 mutation. FLO2 harbors a tetratricopeptide repeat motif, considered to mediate a protein-protein interactions. FLO2 was abundantly expressed in developing seeds coincident with production of storage starch and protein, as well as in leaves, while abundant expression of its homologs was observed only in leaves. The flo2 mutation decreased expression of genes involved in production of storage starch and storage proteins in the endosperm. Differences between cultivars in their responsiveness of FLO2 expression during high-temperature stress indicated that FLO2 may be involved in heat tolerance during seed development. Overexpression of FLO2 enlarged the size of grains significantly. These results suggest that FLO2 plays a pivotal regulatory role in rice grain size and starch quality by affecting storage substance accumulation in the endosperm.


Assuntos
Endosperma/crescimento & desenvolvimento , Oryza/genética , Proteínas de Armazenamento de Sementes/metabolismo , Amido/análise , Amilopectina/análise , Amilose/análise , Mapeamento Cromossômico , Clonagem Molecular , DNA de Plantas/genética , Regulação da Expressão Gênica de Plantas , Teste de Complementação Genética , Glucanos/análise , Temperatura Alta , Dados de Sequência Molecular , Mutação , Oryza/metabolismo , Filogenia , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Proteínas de Armazenamento de Sementes/genética
11.
Genes Dev ; 23(4): 512-21, 2009 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-19196655

RESUMO

The gaseous plant hormone ethylene can trigger myriad physiological and morphological responses in plants. While many ethylene signaling pathway components have been identified and characterized, little is known about the function of the integral membrane protein ETHYLENE-INSENSITIVE2 (EIN2), a central regulator of all ethylene responses. Here, we demonstrate that Arabidopsis thaliana EIN2 is a protein with a short half-life that undergoes rapid proteasome-mediated protein turnover. Moreover, EIN2 protein accumulation is positively regulated by ethylene. We identified two F-box proteins, EIN2 TARGETING PROTEIN1 (ETP1) and EIN2 TARGETING PROTEIN2 (ETP2), that interact with the EIN2 C-terminal domain (EIN2-CEND), which is highly conserved and sufficient to activate most ethylene responses. Overexpression of ETP1 or ETP2 disrupts EIN2 protein accumulation, and these plants manifest a strong ethylene-insensitive phenotype. Furthermore, knocking down the levels of both ETP1 and ETP2 mRNAs using an artificial microRNA (amiRNA) leads to accumulation of EIN2 protein, resulting in plants that display constitutive ethylene response phenotypes. Finally, ethylene down-regulates ETP1 and ETP2 proteins, impairing their ability to interact with EIN2. Thus, these studies reveal that a complex interplay between ethylene, the regulation of ETP1/ETP2 F-box proteins, and subsequent targeting and degradation of EIN2 is essential for triggering ethylene responses in plants.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Etilenos/metabolismo , Proteínas F-Box/metabolismo , Receptores de Superfície Celular/metabolismo , Sequência de Aminoácidos , Proteínas de Arabidopsis/química , Proteínas F-Box/química , Expressão Gênica , Técnicas de Silenciamento de Genes , Meia-Vida , Dados de Sequência Molecular , Alinhamento de Sequência
12.
Nat Methods ; 6(1): 39-46, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19116613

RESUMO

High-quality datasets are needed to understand how global and local properties of protein-protein interaction, or 'interactome', networks relate to biological mechanisms, and to guide research on individual proteins. In an evaluation of existing curation of protein interaction experiments reported in the literature, we found that curation can be error-prone and possibly of lower quality than commonly assumed.


Assuntos
Bases de Dados de Proteínas , Proteínas/metabolismo , Animais , Bases de Dados Factuais , Humanos , Ligação Proteica , Proteínas/análise , Proteínas/química , Reprodutibilidade dos Testes , Projetos de Pesquisa
13.
J Plant Physiol ; 166(7): 750-61, 2009 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-19046615

RESUMO

Apomixis is an intriguing asexual mode of reproduction, because it produces maternal clones that permit vegetative reproduction through seeds. Guineagrass (Panicum maximum) has both facultative aposporous apomixis and obligate sexual modes of reproduction. Despite the importance of apomixis in guineagrass, expressed sequence tags (ESTs) for this condition have not been studied in this species. We constructed a guineagrass cDNA library from two aposporous strains, Ku5954 and GM64-3A, and utilized them as microarray probes. To find genes uniquely expressed in the immature pistils of apomicts, we performed a microarray analysis using target RNA from another apomict, OKI64. Of the 4608 probes in the microarray, only 394 showed clear gene expression in the immature pistils. Of the 394 expressed probes, 196 were successfully sequenced. Of these, 181 had significant homology with other species, including 10 ESTs with matches in a pistil cDNA library from another aposporous species, Cenchrus ciliaris. Of the remaining ESTs, three showed significant homology only with animal database sequences and the other 12 ESTs showed no homology with any previously registered sequence. In reverse-transcriptase PCR and real-time quantitative PCR, nine ESTs reliably detected ovary-specific gene expression. Of these, three revealed aposporous ovary-specific genes expressed in the early developmental stage, suggesting that these could be apomixis-related genes.


Assuntos
Etiquetas de Sequências Expressas , Panicum/genética , Panicum/fisiologia , Flores/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Biblioteca Gênica , Folhas de Planta/genética , Reprodução/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa
14.
Plant J ; 53(4): 636-44, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18269573

RESUMO

The recent explosion in available genome sequence data has ushered in an era in which analysis of a whole genome can be performed in a single experiment. While DNA microarrays have long been the established technology for measuring gene expression levels, standard expression arrays use relatively few probes for each gene and are typically biased toward known and predicted gene structures. Recently, with the availability of complete genome sequences for many organisms, very-high-density oligonucleotide-based microarrays that span the entire genome have emerged as the preferred platform for genomic analysis. Whole-genome tiling microarrays can be employed for a myriad of purposes, including empirical annotation of the transcriptome, chromatin immunoprecipitation-chip studies, analysis of alternative RNA splicing, characterization of the methylation state of cytosine bases throughout a genome (methylome), and DNA polymorphism discovery. Here, we review several applications of whole-genome technology to obtain a variety of genomic-scale information in plants.


Assuntos
Regulação da Expressão Gênica de Plantas , Genoma de Planta/genética , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Processamento Alternativo , Imunoprecipitação da Cromatina , Modelos Genéticos , Transcrição Gênica
15.
Cell ; 131(7): 1340-53, 2007 Dec 28.
Artigo em Inglês | MEDLINE | ID: mdl-18160042

RESUMO

The exosome complex plays a central and essential role in RNA metabolism. However, comprehensive studies of exosome substrates and functional analyses of its subunits are lacking. Here, we demonstrate that as opposed to yeast and metazoans the plant exosome core possesses an unanticipated functional plasticity and present a genome-wide atlas of Arabidopsis exosome targets. Additionally, our study provides evidence for widespread polyadenylation- and exosome-mediated RNA quality control in plants, reveals unexpected aspects of stable structural RNA metabolism, and uncovers numerous novel exosome substrates. These include a select subset of mRNAs, miRNA processing intermediates, and hundreds of noncoding RNAs, the vast majority of which have not been previously described and belong to a layer of the transcriptome that can only be visualized upon inhibition of exosome activity. These first genome-wide maps of exosome substrates will aid in illuminating new fundamental components and regulatory mechanisms of eukaryotic transcriptomes.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Mapeamento Cromossômico , Exorribonucleases/metabolismo , Perfilação da Expressão Gênica , Plantas Geneticamente Modificadas/metabolismo , Proteômica , RNA/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Mapeamento Cromossômico/métodos , Exorribonucleases/genética , Complexo Multienzimático de Ribonucleases do Exossomo , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Genótipo , MicroRNAs/metabolismo , Dados de Sequência Molecular , Mutação , Proteínas Nucleares/metabolismo , Conformação de Ácido Nucleico , Análise de Sequência com Séries de Oligonucleotídeos , Mapeamento de Peptídeos , Fenótipo , Proteômica/métodos , RNA/química , Interferência de RNA , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , RNA não Traduzido/metabolismo , Espectrometria de Massas em Tandem
16.
Curr Opin Plant Biol ; 10(5): 534-42, 2007 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17703988

RESUMO

With the availability of complete genome sequences for a growing number of organisms, high-throughput methods for gene annotation and analysis of genome dynamics are needed. The application of whole-genome tiling microarrays for studies of global gene expression is providing a more unbiased view of the transcriptional activity within genomes. For example, this approach has led to the identification and isolation of many novel non-protein-coding RNAs (ncRNAs), which have been suggested to comprise a major component of the transcriptome that have novel functions involved in epigenetic regulation of the genome. Additionally, tiling arrays have been recently applied to the study of histone modifications and methylation of cytosine bases (DNA methylation). Surprisingly, recent studies combining the analysis of gene expression (transcriptome) and DNA methylation (methylome) using whole-genome tiling arrays revealed that DNA methylation regulates the expression levels of many ncRNAs. Further capture and integration of additional types of genome-wide data sets will help to illuminate additional hidden features of the dynamic genomic landscape that are regulated by both genetic and epigenetic pathways in plants.


Assuntos
Genoma de Planta , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Plantas/genética , Metilação de DNA , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas
17.
Biosci Biotechnol Biochem ; 70(10): 2410-9, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17031029

RESUMO

Transcripts of thionin genes encoding antimicrobial peptides were present at a high level in rice coleoptiles just after germination, and decreased to an undetectable level after about 3 d, but this decline was suppressed by co-treatment with gibberellic acid (GA(3)) and brassinolide (BL). The temporal expression patterns of key enzyme genes for the biosyntheses of gibberellins (GAs) and brassinosteroids (BRs) were correlated with the fluctuation of thionin mRNAs. Jasmonic acid (JA) replaced the effect of GA3 and BL, and its change in endogenous level was parallel to that of the thionin genes. These results strongly suggest that thionin gene expression was positively regulated by JA, whose endogenous level was synergistically regulated by GAs and BRs. In contrast, thionin gene expression in etiolated seedlings remained high while the endogenous level of JA was low, suggesting the presence of another signaling pathway in the dark to maintain the thionin level.


Assuntos
Peptídeos Catiônicos Antimicrobianos/genética , Colestanóis/metabolismo , Regulação da Expressão Gênica de Plantas , Giberelinas/biossíntese , Proteínas de Plantas/genética , Esteroides Heterocíclicos/metabolismo , Brassinosteroides , Colestanóis/farmacologia , Ciclopentanos/farmacologia , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Giberelinas/farmacologia , Oryza , Oxilipinas , RNA Mensageiro , Plântula/genética , Esteroides Heterocíclicos/farmacologia
18.
Cell ; 126(6): 1189-201, 2006 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-16949657

RESUMO

Cytosine methylation is important for transposon silencing and epigenetic regulation of endogenous genes, although the extent to which this DNA modification functions to regulate the genome is still unknown. Here we report the first comprehensive DNA methylation map of an entire genome, at 35 base pair resolution, using the flowering plant Arabidopsis thaliana as a model. We find that pericentromeric heterochromatin, repetitive sequences, and regions producing small interfering RNAs are heavily methylated. Unexpectedly, over one-third of expressed genes contain methylation within transcribed regions, whereas only approximately 5% of genes show methylation within promoter regions. Interestingly, genes methylated in transcribed regions are highly expressed and constitutively active, whereas promoter-methylated genes show a greater degree of tissue-specific expression. Whole-genome tiling-array transcriptional profiling of DNA methyltransferase null mutants identified hundreds of genes and intergenic noncoding RNAs with altered expression levels, many of which may be epigenetically controlled by DNA methylation.


Assuntos
Arabidopsis/genética , Metilação de DNA , DNA de Plantas/genética , Genes de Plantas/genética , Genoma de Planta/genética , Arabidopsis/metabolismo , Mapeamento Cromossômico/métodos , Metilases de Modificação do DNA/genética , Metilases de Modificação do DNA/metabolismo , DNA de Plantas/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas/genética , Inativação Gênica/fisiologia , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Regiões Promotoras Genéticas/genética , RNA Interferente Pequeno/genética , Transcrição Gênica/genética
19.
Plant J ; 48(1): 85-97, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16972867

RESUMO

Mugineic acid family phytosiderophores (MAs) are metal chelators that are produced in graminaceous plants in response to iron (Fe) deficiency, but current evidence regarding secretion of MAs during zinc (Zn) deficiency is contradictory. Our studies using HPLC analysis showed that Zn deficiency induces the synthesis and secretion of MAs in barley plants. The levels of the HvNAS1, HvNAAT-A, HvNAAT-B, HvIDS2 and HvIDS3 transcripts, which encode the enzymes involved in the synthesis of MAs, were increased in Zn-deficient roots. Studies of the genes involved in the methionine cycle using microarray analysis showed that the transcripts of these genes were increased in both Zn-deficient and Fe-deficient barley roots, probably allowing the plant to meet its demand for methionine, a precursor in the synthesis of MAs. In addition, HvNAAT-B transcripts were detected in Zn-deficient shoots, but not in those that were deficient in Fe. Increased synthesis of MAs in Zn-deficient barley was not due to a deficiency of Fe, because Zn-deficient barley accumulated more Fe than did the control plants, ferritin transcripts were increased in Zn-deficient plants, and Zn deficiency promoted Fe transport from root to shoot. Moreover, analysis using the positron-emitting tracer imaging system (PETIS) confirmed that more 62Zn(II)-MAs than 62Zn2+ were absorbed by the roots of Zn-deficient barley plants. These data suggest that the increased biosynthesis and secretion of MAs arising from a shortage of Zn are not due to an induced Fe deficiency, and that secreted MAs are effective in absorbing Zn from the soil.


Assuntos
Ácido Azetidinocarboxílico/análogos & derivados , Quelantes/metabolismo , Hordeum/metabolismo , Zinco/metabolismo , Ácido Azetidinocarboxílico/química , Ácido Azetidinocarboxílico/metabolismo , Transporte Biológico , Cromatografia Líquida de Alta Pressão , Ferritinas/genética , Ferritinas/metabolismo , Perfilação da Expressão Gênica , Hordeum/genética , Hordeum/crescimento & desenvolvimento , Ferro/análise , Ferro/metabolismo , Metionina/química , Metionina/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Brotos de Planta/genética , Brotos de Planta/crescimento & desenvolvimento , Brotos de Planta/metabolismo , Tomografia por Emissão de Pósitrons , RNA Mensageiro/metabolismo , Regulação para Cima , Zinco/análise
20.
J Exp Bot ; 57(9): 2049-59, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16720613

RESUMO

Plants have developed several strategies for coping with phosphorus (P) deficiency. However, the details of the regulation of gene expression of adaptations to low P are still unclear. Using a cDNA microarray, transcriptomic analyses were carried out of the rice genes regulated by P deficiency and P re-supply to P-deficient plants. The OsPI1 gene, which was isolated as the most significant up-regulated gene under -P conditions, was also the most significant down-regulated gene following P re-supply. Many starch metabolism-related genes, as well as several genes for P(i)-liberating enzymes, were up-regulated by -P treatment, suggesting a homeostatic contribution to the P(i) concentration in leaf tissues. mRNAs for glucanases were also induced by P re-supply: these are suspected to play a role in loosening the cell wall compounds. Most of the genes up-regulated by -P treatment were down-regulated by P re-supply, suggesting that their responses were specific to -P conditions. Conversely, the number of genes up-regulated by P re-supply was also larger following P re-supply than in the -P condition. It is proposed that the genes up-regulated by P re-supply play an important role in P acquisition by P-deficient plants.


Assuntos
Oryza/metabolismo , Fósforo/metabolismo , Folhas de Planta/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Análise de Sequência com Séries de Oligonucleotídeos
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