RESUMO
Land plants evolved from charophytic algae, among which Charophyceae possess the most complex body plans. We present the genome of Chara braunii; comparison of the genome to those of land plants identified evolutionary novelties for plant terrestrialization and land plant heritage genes. C. braunii employs unique xylan synthases for cell wall biosynthesis, a phragmoplast (cell separation) mechanism similar to that of land plants, and many phytohormones. C. braunii plastids are controlled via land-plant-like retrograde signaling, and transcriptional regulation is more elaborate than in other algae. The morphological complexity of this organism may result from expanded gene families, with three cases of particular note: genes effecting tolerance to reactive oxygen species (ROS), LysM receptor-like kinases, and transcription factors (TFs). Transcriptomic analysis of sexual reproductive structures reveals intricate control by TFs, activity of the ROS gene network, and the ancestral use of plant-like storage and stress protection proteins in the zygote.
Assuntos
Chara/genética , Genoma de Planta , Evolução Biológica , Parede Celular/metabolismo , Chara/crescimento & desenvolvimento , Embriófitas/genética , Redes Reguladoras de Genes , Pentosiltransferases/genética , Reguladores de Crescimento de Plantas/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , TranscriptomaRESUMO
Cultivated oat (Avena sativa L.) is an allohexaploid (AACCDD, 2n = 6x = 42) thought to have been domesticated more than 3,000 years ago while growing as a weed in wheat, emmer and barley fields in Anatolia1,2. Oat has a low carbon footprint, substantial health benefits and the potential to replace animal-based food products. However, the lack of a fully annotated reference genome has hampered efforts to deconvolute its complex evolutionary history and functional gene dynamics. Here we present a high-quality reference genome of A. sativa and close relatives of its diploid (Avena longiglumis, AA, 2n = 14) and tetraploid (Avena insularis, CCDD, 2n = 4x = 28) progenitors. We reveal the mosaic structure of the oat genome, trace large-scale genomic reorganizations in the polyploidization history of oat and illustrate a breeding barrier associated with the genome architecture of oat. We showcase detailed analyses of gene families implicated in human health and nutrition, which adds to the evidence supporting oat safety in gluten-free diets, and we perform mapping-by-sequencing of an agronomic trait related to water-use efficiency. This resource for the Avena genus will help to leverage knowledge from other cereal genomes, improve understanding of basic oat biology and accelerate genomics-assisted breeding and reanalysis of quantitative trait studies.
Assuntos
Avena , Grão Comestível , Genoma de Planta , Avena/genética , Diploide , Grão Comestível/genética , Genoma de Planta/genética , Mosaicismo , Melhoramento Vegetal , TetraploidiaRESUMO
Plants are essential for life and are extremely diverse organisms with unique molecular capabilities1. Here we present a quantitative atlas of the transcriptomes, proteomes and phosphoproteomes of 30 tissues of the model plant Arabidopsis thaliana. Our analysis provides initial answers to how many genes exist as proteins (more than 18,000), where they are expressed, in which approximate quantities (a dynamic range of more than six orders of magnitude) and to what extent they are phosphorylated (over 43,000 sites). We present examples of how the data may be used, such as to discover proteins that are translated from short open-reading frames, to uncover sequence motifs that are involved in the regulation of protein production, and to identify tissue-specific protein complexes or phosphorylation-mediated signalling events. Interactive access to this resource for the plant community is provided by the ProteomicsDB and ATHENA databases, which include powerful bioinformatics tools to explore and characterize Arabidopsis proteins, their modifications and interactions.
Assuntos
Proteínas de Arabidopsis/análise , Proteínas de Arabidopsis/química , Arabidopsis/química , Espectrometria de Massas , Proteoma/análise , Proteoma/química , Proteômica , Motivos de Aminoácidos , Arabidopsis/anatomia & histologia , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/biossíntese , Proteínas de Arabidopsis/genética , Bases de Dados de Proteínas , Conjuntos de Dados como Assunto , Regulação da Expressão Gênica de Plantas , Anotação de Sequência Molecular , Fases de Leitura Aberta , Especificidade de Órgãos , Fosfoproteínas/análise , Fosfoproteínas/química , Fosfoproteínas/genética , Fosforilação , Proteoma/biossíntese , Proteoma/genética , RNA Mensageiro/análise , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , TranscriptomaRESUMO
Genetic diversity is key to crop improvement. Owing to pervasive genomic structural variation, a single reference genome assembly cannot capture the full complement of sequence diversity of a crop species (known as the 'pan-genome'1). Multiple high-quality sequence assemblies are an indispensable component of a pan-genome infrastructure. Barley (Hordeum vulgare L.) is an important cereal crop with a long history of cultivation that is adapted to a wide range of agro-climatic conditions2. Here we report the construction of chromosome-scale sequence assemblies for the genotypes of 20 varieties of barley-comprising landraces, cultivars and a wild barley-that were selected as representatives of global barley diversity. We catalogued genomic presence/absence variants and explored the use of structural variants for quantitative genetic analysis through whole-genome shotgun sequencing of 300 gene bank accessions. We discovered abundant large inversion polymorphisms and analysed in detail two inversions that are frequently found in current elite barley germplasm; one is probably the product of mutation breeding and the other is tightly linked to a locus that is involved in the expansion of geographical range. This first-generation barley pan-genome makes previously hidden genetic variation accessible to genetic studies and breeding.
Assuntos
Cromossomos de Plantas/genética , Genoma de Planta/genética , Hordeum/genética , Internacionalidade , Mutação , Melhoramento Vegetal , Inversão Cromossômica/genética , Mapeamento Cromossômico , Loci Gênicos/genética , Genótipo , Hordeum/classificação , Polimorfismo Genético/genética , Padrões de Referência , Banco de Sementes , Inversão de Sequência , Sequenciamento Completo do GenomaRESUMO
Breeding has increasingly altered the genetics of crop plants since the domestication of their wild progenitors. It is postulated that the genetic diversity of elite wheat breeding pools is too narrow to cope with future challenges. In contrast, plant genetic resources (PGRs) of wheat stored in genebanks are valuable sources of unexploited genetic diversity. Therefore, to ensure breeding progress in the future, it is of prime importance to identify the useful allelic diversity available in PGRs and to transfer it into elite breeding pools. Here, a diverse collection consisting of modern winter wheat cultivars and genebank accessions was investigated based on reduced-representation genomic sequencing and an iSelect single nucleotide polymorphism (SNP) chip array. Analyses of these datasets provided detailed insights into population structure, levels of genetic diversity, sources of new allelic diversity, and genomic regions affected by breeding activities. We identified 57 regions representing genomic signatures of selection and 827 regions representing private alleles associated exclusively with genebank accessions. The presence of known functional wheat genes, quantitative trait loci, and large chromosomal modifications, i.e., introgressions from wheat wild relatives, provided initial evidence for putative traits associated within these identified regions. These findings were supported by the results of ontology enrichment analyses. The results reported here will stimulate further research and promote breeding in the future by allowing for the targeted introduction of novel allelic diversity into elite wheat breeding pools.
Assuntos
Pão , Triticum , Triticum/genética , Alelos , Melhoramento Vegetal , Genoma de Planta/genética , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
In July 2022, a case of paralytic poliomyelitis resulting from infection with vaccine-derived poliovirus (VDPV) type 2 (VDPV2)§ was confirmed in an unvaccinated adult resident of Rockland County, New York (1). As of August 10, 2022, poliovirus type 2 (PV2)¶ genetically linked to this VDPV2 had been detected in wastewater** in Rockland County and neighboring Orange County (1). This report describes the results of additional poliovirus testing of wastewater samples collected during March 9-October 11, 2022, and tested as of October 20, 2022, from 48 sewersheds (the community area served by a wastewater collection system) serving parts of Rockland County and 12 surrounding counties. Among 1,076 wastewater samples collected, 89 (8.3%) from 10 sewersheds tested positive for PV2. As part of a broad epidemiologic investigation, wastewater testing can provide information about where poliovirus might be circulating in a community in which a paralytic case has been identified; however, the most important public health actions for preventing paralytic poliomyelitis in the United States remain ongoing case detection through national acute flaccid myelitis (AFM) surveillance and improving vaccination coverage in undervaccinated communities. Although most persons in the United States are sufficiently immunized, unvaccinated or undervaccinated persons living or working in Kings, Orange, Queens, Rockland, or Sullivan counties, New York should complete the polio vaccination series as soon as possible.
Assuntos
Poliomielite , Vacina Antipólio Oral , Poliovirus , Adulto , Humanos , New York/epidemiologia , Poliomielite/diagnóstico , Poliomielite/epidemiologia , Poliomielite/prevenção & controle , Poliovirus/genética , Vacina Antipólio Oral/efeitos adversos , Estados Unidos , Águas ResiduáriasRESUMO
Wheat has been domesticated into a large number of agricultural environments and has the ability to adapt to diverse environments. To understand this process, we survey genotype, repeat content, and DNA methylation across a bread wheat landrace collection representing global genetic diversity. We identify independent variation in methylation, genotype, and transposon copy number. We show that these, so far unexploited, sources of variation have had a significant impact on the wheat genome and that ancestral methylation states become preferentially "hard coded" as single nucleotide polymorphisms (SNPs) via 5-methylcytosine deamination. These mechanisms also drive local adaption, impacting important traits such as heading date and salt tolerance. Methylation and transposon diversity could therefore be used alongside SNP-based markers for breeding.
Assuntos
Adaptação Fisiológica/genética , Variação Genética , Poliploidia , Triticum/genética , Metilação de DNA , Elementos de DNA Transponíveis/genéticaRESUMO
Next-generation sequencing technologies have made it possible to carry out transcriptome analysis at the single-cell level. Single-cell RNA-sequencing (scRNA-seq) data provide insights into cellular dynamics, including intercellular heterogeneity as well as inter- and intra-cellular fluctuations in gene expression that cannot be studied using populations of cells. The utilization of scRNA-seq is, however, restricted to cell types that can be isolated from their original tissues, and it can be difficult to obtain precise positional information for these cells in situ. Here, we established single cell-digital gene expression (1cell-DGE), a method of scRNA-seq that uses micromanipulation to extract the contents of individual living cells in intact tissue while recording their positional information. With 1cell-DGE, we could detect differentially expressed genes (DEGs) during the reprogramming of leaf cells of the moss Physcomitrella patens, identifying 6382 DEGs between cells at 0 and 24 h after excision. Furthermore, we identified a subpopulation of reprogramming cells based on their pseudotimes, which were calculated using transcriptome profiles at 24 h. 1cell-DGE with microcapillary manipulation can be used to analyze the gene expression of individual cells without detaching them from their tightly associated tissues, enabling us to retain positional information and investigate cell-cell interactions.
Assuntos
Bryopsida/genética , Reprogramação Celular/genética , Perfilação da Expressão Gênica/métodos , Análise de Célula Única/métodos , Folhas de Planta/genética , Análise de Sequência de RNA/métodos , Software , Transcriptoma/genéticaRESUMO
Nonsense-mediated mRNA decay (NMD) is important for RNA quality control and gene regulation in eukaryotes. NMD targets aberrant transcripts for decay and also directly influences the abundance of non-aberrant transcripts. In animals, the SMG1 kinase plays an essential role in NMD by phosphorylating the core NMD factor UPF1. Despite SMG1 being ubiquitous throughout the plant kingdom, little is known about its function, probably because SMG1 is atypically absent from the genome of the model plant, Arabidopsis thaliana. By combining our previously established SMG1 knockout in moss with transcriptome-wide analysis, we reveal the range of processes involving SMG1 in plants. Machine learning assisted analysis suggests that 32% of multi-isoform genes produce NMD-targeted transcripts and that splice junctions downstream of a stop codon act as the major determinant of NMD targeting. Furthermore, we suggest that SMG1 is involved in other quality control pathways, affecting DNA repair and the unfolded protein response, in addition to its role in mRNA quality control. Consistent with this, smg1 plants have increased susceptibility to DNA damage, but increased tolerance to unfolded protein inducing agents. The potential involvement of SMG1 in RNA, DNA and protein quality control has major implications for the study of these processes in plants.
Assuntos
Bryopsida/enzimologia , Bryopsida/genética , Degradação do RNAm Mediada por Códon sem Sentido , Fosfotransferases/fisiologia , Proteínas de Plantas/fisiologia , Regiões 3' não Traduzidas , Bryopsida/metabolismo , Dano ao DNA , Expressão Gênica , Mutação , Fosfotransferases/genética , Proteínas de Plantas/genética , Resposta a Proteínas não DobradasRESUMO
Health care organizations that deliver health care to a population are a major constituent of health care systems in many countries. Although some of the components of a health care organization may function as a simple or complicated system where interventions would yield the expected results, many parts of it function as complex systems within the organization. As such, is not always possible to predict changes or the effects of interventions on these systems due to their complex nature. Appreciation of the complex nature of health care systems and characteristics of these complex systems and their networks are necessary to manage health care organizations and changes in health care. This article described different types of systems and discusses why health care organizations are considered complex adaptive systems.
Assuntos
Atenção à Saúde/organização & administração , Inovação Organizacional , Humanos , Dinâmica não Linear , Análise de SistemasRESUMO
High-throughput RNA sequencing (RNA-seq) has recently become the method of choice to define and analyze transcriptomes. For the model moss Physcomitrella patens, although this method has been used to help analyze specific perturbations, no overall reference dataset has yet been established. In the framework of the Gene Atlas project, the Joint Genome Institute selected P. patens as a flagship genome, opening the way to generate the first comprehensive transcriptome dataset for this moss. The first round of sequencing described here is composed of 99 independent libraries spanning 34 different developmental stages and conditions. Upon dataset quality control and processing through read mapping, 28 509 of the 34 361 v3.3 gene models (83%) were detected to be expressed across the samples. Differentially expressed genes (DEGs) were calculated across the dataset to permit perturbation comparisons between conditions. The analysis of the three most distinct and abundant P. patens growth stages - protonema, gametophore and sporophyte - allowed us to define both general transcriptional patterns and stage-specific transcripts. As an example of variation of physico-chemical growth conditions, we detail here the impact of ammonium supplementation under standard growth conditions on the protonemal transcriptome. Finally, the cooperative nature of this project allowed us to analyze inter-laboratory variation, as 13 different laboratories around the world provided samples. We compare differences in the replication of experiments in a single laboratory and between different laboratories.
Assuntos
Bryopsida/genética , Conjuntos de Dados como Assunto , Genes de Plantas/genética , Mapeamento Cromossômico , Genoma de Planta/genética , Sequenciamento de Nucleotídeos em Larga Escala , Transcriptoma/genéticaRESUMO
The draft genome of the moss model, Physcomitrella patens, comprised approximately 2000 unordered scaffolds. In order to enable analyses of genome structure and evolution we generated a chromosome-scale genome assembly using genetic linkage as well as (end) sequencing of long DNA fragments. We find that 57% of the genome comprises transposable elements (TEs), some of which may be actively transposing during the life cycle. Unlike in flowering plant genomes, gene- and TE-rich regions show an overall even distribution along the chromosomes. However, the chromosomes are mono-centric with peaks of a class of Copia elements potentially coinciding with centromeres. Gene body methylation is evident in 5.7% of the protein-coding genes, typically coinciding with low GC and low expression. Some giant virus insertions are transcriptionally active and might protect gametes from viral infection via siRNA mediated silencing. Structure-based detection methods show that the genome evolved via two rounds of whole genome duplications (WGDs), apparently common in mosses but not in liverworts and hornworts. Several hundred genes are present in colinear regions conserved since the last common ancestor of plants. These syntenic regions are enriched for functions related to plant-specific cell growth and tissue organization. The P. patens genome lacks the TE-rich pericentromeric and gene-rich distal regions typical for most flowering plant genomes. More non-seed plant genomes are needed to unravel how plant genomes evolve, and to understand whether the P. patens genome structure is typical for mosses or bryophytes.
Assuntos
Evolução Biológica , Bryopsida/genética , Cromossomos de Plantas , Genoma de Planta , Centrômero , Cromatina/genética , Metilação de DNA , Elementos de DNA Transponíveis , Variação Genética , Polimorfismo de Nucleotídeo Único , Recombinação Genética , SinteniaRESUMO
The anatomically simple plants that first colonized land must have acquired molecular and biochemical adaptations to drought stress. Abscisic acid (ABA) coordinates responses leading to desiccation tolerance in all land plants. We identified ABA nonresponsive mutants in the model bryophyte Physcomitrella patens and genotyped a segregating population to map and identify the ABA NON-RESPONSIVE (ANR) gene encoding a modular protein kinase comprising an N-terminal PAS domain, a central EDR domain, and a C-terminal MAPKKK-like domain. anr mutants fail to accumulate dehydration tolerance-associated gene products in response to drought, ABA, or osmotic stress and do not acquire ABA-dependent desiccation tolerance. The crystal structure of the PAS domain, determined to 1.7-Å resolution, shows a conserved PAS-fold that dimerizes through a weak dimerization interface. Targeted mutagenesis of a conserved tryptophan residue within the PAS domain generates plants with ABA nonresponsive growth and strongly attenuated ABA-responsive gene expression, whereas deleting this domain retains a fully ABA-responsive phenotype. ANR orthologs are found in early-diverging land plant lineages and aquatic algae but are absent from more recently diverged vascular plants. We propose that ANR genes represent an ancestral adaptation that enabled drought stress survival of the first terrestrial colonizers but were lost during land plant evolution.
Assuntos
Ácido Abscísico/farmacologia , Bryopsida/efeitos dos fármacos , Bryopsida/metabolismo , Proteínas de Plantas/metabolismo , Bryopsida/genética , Cristalografia por Raios X , Dessecação , Secas , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/genética , Mutação , Pressão Osmótica , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Estrutura Secundária de ProteínaRESUMO
Host cell proteins are inevitable contaminants of biopharmaceuticals. Here, we performed detailed analyses of the host cell proteome of moss ( Physcomitrella patens) bioreactor supernatants using mass spectrometry and subsequent bioinformatics analysis. Distinguishing between the apparent secretome and intracellular contaminants, a complex extracellular proteolytic network including subtilisin-like proteases, metallo-proteases, and aspartic proteases was identified. Knockout of a subtilisin-like protease affected the overall extracellular proteolytic activity. Besides proteases, also secreted protease-inhibiting proteins such as serpins were identified. Further, we confirmed predicted cleavage sites of 40 endogenous signal peptides employing an N-terminomics approach. The present data provide novel aspects to optimize both product stability of recombinant biopharmaceuticals as well as their maturation along the secretory pathway. Data are available via ProteomeXchange with identifier PXD009517.
Assuntos
Ácido Aspártico Proteases/isolamento & purificação , Bryopsida/enzimologia , Metaloproteases/isolamento & purificação , Proteínas de Plantas/isolamento & purificação , Inibidores de Proteases/isolamento & purificação , Serpinas/isolamento & purificação , Subtilisinas/isolamento & purificação , Ácido Aspártico Proteases/classificação , Ácido Aspártico Proteases/genética , Ácido Aspártico Proteases/metabolismo , Reatores Biológicos , Bryopsida/química , Bryopsida/genética , Biologia Computacional , Técnicas de Inativação de Genes , Espectrometria de Massas/métodos , Metaloproteases/classificação , Metaloproteases/genética , Metaloproteases/metabolismo , Proteínas de Plantas/classificação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Inibidores de Proteases/classificação , Inibidores de Proteases/metabolismo , Análise Serial de Proteínas , Proteólise , Serpinas/classificação , Serpinas/genética , Serpinas/metabolismo , Subtilisinas/classificação , Subtilisinas/genética , Subtilisinas/metabolismoRESUMO
Molecular evolutionary studies correlate genomic and phylogenetic information with the emergence of new traits of organisms. These traits are, however, the consequence of dynamic gene networks composed of functional modules, which might not be captured by genomic analyses. Here, we established a method that combines large-scale genomic and phylogenetic data with gene co-expression networks to extensively study the evolutionary make-up of modules in the moss Physcomitrella patens, and in the angiosperms Arabidopsis thaliana and Oryza sativa (rice). We first show that younger genes are less annotated than older genes. By mapping genomic data onto the co-expression networks, we found that genes from the same evolutionary period tend to be connected, whereas old and young genes tend to be disconnected. Consequently, the analysis revealed modules that emerged at a specific time in plant evolution. To uncover the evolutionary relationships of the modules that are conserved across the plant kingdom, we added phylogenetic information that revealed duplication and speciation events on the module level. This combined analysis revealed an independent duplication of cell wall modules in bryophytes and angiosperms, suggesting a parallel evolution of cell wall pathways in land plants. We provide an online tool allowing plant researchers to perform these analyses at http://www.gene2function.de.
Assuntos
Proteínas de Plantas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Bryopsida/genética , Bryopsida/metabolismo , Evolução Molecular , Oryza/genética , Oryza/metabolismo , Filogenia , Proteínas de Plantas/genéticaRESUMO
BACKGROUND: Chronic infection with the neurotropic parasite Toxoplasma gondii has been implicated in the risk for several neuropsychiatric disorders. The mechanisms, by which the parasite may alter neural function and behavior of the host, are not yet understood completely. METHODS: Here, a novel proteomic approach using mass spectrometry was employed to investigate the alterations in synaptic protein composition in a murine model of chronic toxoplasmosis. In a candidate-based strategy, immunoblot analysis and immunohistochemistry were applied to investigate the expression levels of key synaptic proteins in glutamatergic signaling. RESULTS: A comparison of the synaptosomal protein composition revealed distinct changes upon infection, with multiple proteins such as EAAT2, Shank3, AMPA receptor, and NMDA receptor subunits being downregulated, whereas inflammation-related proteins showed an upregulation. Treatment with the antiparasitic agent sulfadiazine strongly reduced tachyzoite levels and diminished neuroinflammatory mediators. However, in both conditions, a significant number of latent cysts persisted in the brain. Conversely, infection-related alterations of key synaptic protein levels could be partly reversed by the treatment. CONCLUSION: These results provide evidence for profound changes especially in synaptic protein composition in T. gondii-infected mice with a downregulation of pivotal components of glutamatergic neurotransmission. Our results suggest that the detected synaptic alterations are a consequence of the distinct neuroinflammatory milieu caused by the neurotropic parasite.
Assuntos
Encéfalo/metabolismo , Regulação da Expressão Gênica/fisiologia , Sinapses/metabolismo , Sinaptossomos/metabolismo , Toxoplasmose Animal/patologia , Animais , Antiprotozoários/farmacologia , Doença Crônica , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Ácido Glutâmico/metabolismo , Espectrometria de Massas , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Metanálise como Assunto , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteômica , RNA Mensageiro/metabolismo , Sulfadiazina/farmacologia , Sinapses/patologia , Sinaptossomos/efeitos dos fármacos , Espectrometria de Massas em Tandem , Toxoplasma/patogenicidadeRESUMO
Protein arginylation is a posttranslational modification of both N-terminal amino acids of proteins and sidechain carboxylates and can be crucial for viability and physiology in higher eukaryotes. The lack of arginylation causes severe developmental defects in moss, affects the low oxygen response in Arabidopsis thaliana and is embryo lethal in Drosophila and in mice. Although several studies investigated impact and function of the responsible enzyme, the arginyl-tRNA protein transferase (ATE) in plants, identification of arginylated proteins by mass spectrometry was not hitherto achieved. In the present study, we report the identification of targets and interaction partners of ATE in the model plant Physcomitrella patens by mass spectrometry, employing two different immuno-affinity strategies and a recently established transgenic ATE:GUS reporter line (Schuessele et al., 2016 New Phytol. , DOI: 10.1111/nph.13656). Here we use a commercially available antibody against the fused reporter protein (ß-glucuronidase) to pull down ATE and its interacting proteins and validate its in vivo interaction with a class I small heatshock protein via Förster resonance energy transfer (FRET). Additionally, we apply and modify a method that already successfully identified arginylated proteins from mouse proteomes by using custom-made antibodies specific for N-terminal arginine. As a result, we identify four arginylated proteins from Physcomitrella patens with high confidence.Data are available via ProteomeXchange with identifier PXD003228 and PXD003232.
Assuntos
Aminoaciltransferases/metabolismo , Bryopsida/metabolismo , Proteínas de Plantas/metabolismo , Anticorpos/metabolismo , Transferência Ressonante de Energia de Fluorescência , Espectrometria de Massas , Proteínas de Plantas/química , Mapas de Interação de Proteínas , Proteômica/métodosRESUMO
Chronic obstructive pulmonary disease (COPD) is a risk factor of post-operative complications after lung cancer resection. The influence of the "frequent exacerbator (FE)" phenotype (at least three exacerbations per year) is unknown. Postoperative outcomes of frequent exacerbators (POFE) was a prospective observational study of patients with COPD undergoing lung resection for cancer. The inclusion criteria were: age >40 years, FEV1/FVC <70%, non-urgent surgery for lung cancer, filled out self-questionnaires. The primary outcome was assessment of postoperative pulmonary complications (purulent tracheobronchitis, atelectasis, pneumonia, acute respiratory failure, need of mechanical ventilation). Secondary outcomes encompassed the prevalence of the FE phenotype and its impact on postoperative complications. A total of 682 patients were screened from June 2014 to October 2015. 93 patients with COPD were included, 21 (23%) were FE. Postoperative tracheobronchitis, atelectasis pneumonia or respiratory failure (isolated or associated) occurred in 47%, 48%, 26%, and 38% of patients, respectively. Non-invasive and invasive mechanical ventilation were necessary in 4 (4%) and 22 (23%) patients. Purulent tracheobronchitis, pneumonia and hypercapnia (this last requiring noninvasive mechanical ventilation) were more frequent in FE (p = 0.043, 0.042, 0.015); however the number of patients wth at least one respiratory complication was not different (76% vs. 52%, p = 0.056). In all patients, multivariate logistic regression identified two independent factors of postoperative respiratory complications: male sex (OR 10.6 [95% CI 1.97-57.6], p = 0.006) and the FE phenotype (OR 6.33 [1.04-38.39], p = 0.045). Occurrence of postoperative complications in patients with COPD is high. FE phenotype is an independent risk factor.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/cirurgia , Neoplasias Pulmonares/cirurgia , Pneumonectomia , Complicações Pós-Operatórias/epidemiologia , Atelectasia Pulmonar/epidemiologia , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Insuficiência Respiratória/epidemiologia , Infecções Respiratórias/epidemiologia , Idoso , Bronquite/epidemiologia , Carcinoma Pulmonar de Células não Pequenas/complicações , Progressão da Doença , Feminino , Volume Expiratório Forçado , Humanos , Modelos Logísticos , Neoplasias Pulmonares/complicações , Masculino , Pessoa de Meia-Idade , Razão de Chances , Fenótipo , Pneumonia/epidemiologia , Estudos Prospectivos , Doença Pulmonar Obstrutiva Crônica/complicações , Respiração Artificial/estatística & dados numéricos , Insuficiência Respiratória/terapia , Fatores de Risco , Fatores Sexuais , Inquéritos e Questionários , Traqueíte/epidemiologia , Capacidade VitalRESUMO
Packaging of eukaryotic DNA largely depends on histone modifications that affect the accessibility of DNA to transcriptional regulators, thus controlling gene expression. The Polycomb group (PcG) chromatin remodeling complex deposits a methyl group on lysine 27 of histone 3 leading to repressed gene expression. Plants encode homologs of the Enhancer of zeste (E(z)), a component of the PcG complex from Drosophila, one of which is a SET domain protein designated CURLY LEAF (CLF). Although this SET domain protein exhibits a strong correlation with the presence of the H3K27me3 mark in plants, the methyl-transferase activity and specificity of its SET domain have not been directly tested in-vivo. Using the evolutionary early-diverged land plant model species Physcomitrella patens we show that abolishment of a single copy gene PpCLF, as well as an additional member of the PcG complex, FERTILIZATION-INDEPENDENT ENDOSPERM (PpFIE), results in a specific loss of tri-methylation of H3K27. Using site-directed mutagenesis of key residues, we revealed that H3K27 tri-methylation is mediated by the SET domain of the CLF protein. Moreover, the abolishment of H3K27me3 led to enhanced expression of transcription factor genes. This in turn led to the development of fertilization-independent sporophyte-like structures, as observed in PpCLF and PpFIE null mutants. Overall, our results demonstrate the role of PpCLF as a SET protein in tri-methylation of H3K27 in-vivo and the importance of this modification in regulating the expression of transcription factor genes involved in developmental programs of P. patens.
Assuntos
Bryopsida/crescimento & desenvolvimento , Bryopsida/genética , Histona-Lisina N-Metiltransferase/fisiologia , Histonas/metabolismo , Proteínas do Grupo Polycomb/fisiologia , Sequência de Aminoácidos , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/química , Proteínas de Homeodomínio/fisiologia , Lisina/metabolismo , Metilação , Dados de Sequência Molecular , Folhas de Planta/genética , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas , Homologia de Sequência de AminoácidosRESUMO
The importance of the arginyl-tRNA protein transferase (ATE), the enzyme mediating post-translation arginylation of proteins in the N-end rule degradation (NERD) pathway of protein stability, was analysed in Physcomitrella patens and compared to its known functions in other eukaryotes. We characterize ATE:GUS reporter lines as well as ATE mutants in P. patens to study the impact and function of arginylation on moss development and physiology. ATE protein abundance is spatially and temporally regulated in P. patens by hormones and light and is highly abundant in meristematic cells. Further, the amount of ATE transcript is regulated during abscisic acid signalling and downstream of auxin signalling. Loss-of-function mutants exhibit defects at various levels, most severely in developing gametophores, in chloroplast starch accumulation and senescence. Thus, arginylation is necessary for moss gametophyte development, in contrast to the situation in flowering plants. Our analysis further substantiates the conservation of the N-end rule pathway components in land plants and highlights lineage-specific features. We introduce moss as a model system to characterize the role of the NERD pathway as an additional layer of complexity in eukaryotic development.