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Dissociabilidade das funções de inibição da expansão celular e de alcalinização do peptídeo AtRALF1 e identificação dos aminoácidos determinantes da atividade de alcalinização; The dissociability of the root growth inhibition and extracellular alkalinization activities of the AtRALF1 peptide and the identification of the essential amino acids for the alkalinization activity

Ceciliato, Paulo Henrique de Oliveira
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 22/05/2015 PT
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26.87%
RALFs são peptídeos hormonais de aproximadamente 5kD que regulam negativamente o alongamento celular. Dentre suas atividades biológicas, a alcalinização do meio extracelular e a inibição do alongamento celular são tidas como associadas, pois a acidificação do meio extracelular é necessária para a expansão celular. A atividade de alcalinização e a de inibição do crescimento são medidas em dois ensaios distintos: o ensaio de alcalinização do meio extracelular de células em suspensão e o ensaio do crescimento de raiz primária de plantas jovens. Buscando-se fazer ambas as avaliações da inibição da expansão celular e da alcalinização em um único modelo de estudo, tomou-se como medida da expansão celular o volume das células decantadas (VCD). Quando o tampão MES ou o pré-tratamento com "Fusicoccin" foi utilizado para se atenuar o efeito de alcalinização do AtRALF1, verificou-se que o efeito de inibição do alongamento celular não sofreu alteração. Em arabidopsis são encontrados nove peptídeos RALF, que apresentam alta similaridade com o RALF original de tabaco. Com exceção do AtRALF4, todos são capazes de alcalinizar o meio extracelular e inibir raízes. A comparação da estrutura primária do AtRALF4 com os demais RALFs mostra que poucos resíduos são distintos entre eles...

Superexpressão do gene codificante do peptídeo AtPep1 em A.Thaliana visando a obtenção de resistência à isolados de diferentes espécies do gênero Pythium; Overexpression of the gene that codes for the peptide atpep1 in A. Thaliana in order to obtain resistance to isolates of different species of the genus pythium

Trivilin, Ana Paula
Fonte: Universidade Federal do Rio Grande do Sul Publicador: Universidade Federal do Rio Grande do Sul
Tipo: Dissertação Formato: application/pdf
POR
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26.84%
As plantas estão expostas a uma gama de patógenos que utilizam diversas estratégias para infectar seus hospedeiros. Em resposta, as plantas expressam diferentes mecanismos de defesa, onde ocorre o reconhecimento de moléculas conservadas do patógeno por receptores específicos das plantas, desencadeando as respostas de defesa. A descoberta de sinais endógenos e exógenos que regulam a expressão de genes de defesa em Arabidopsis thaliana tem auxiliado a compreensão dos mecanismos de defesa. O desenvolvimento de uma estratégia de superexpressão para avaliação do papel do peptídeo endógeno AtPep1 na resistência a isolados de diferentes espécies do gênero Pythium é importante para a compreensão do mecanismo de defesa e a futura utilização do mesmo em cultivares resistentes ao patógeno. Nesse sentido, foram realizados experimentos que consistiram da inoculação de A. thaliana com isolados das espécies de P. graminicola, P. inflatum, P. deliense e P. ultimum. As plantas foram avaliadas quanto à presença de sintomas da doença. Paralelamente, o gene codificante do peptídeo AtPep1, PROPEP1, foi isolado de A. thaliana, ligado nos vetores binários pGSA1427 ou pEGAD, e transformados em Agrobacterium tumefaciens. Células de A. tumefaciens contendo os plasmídeos com e sem PROPEP1 foram usadas para transformação floral de A. thaliana. As plantas transformadas foram selecionadas pela presença do gene que confere tolerância ao herbicida glufosinato de amônio e...

An endogenous peptide signal in Arabidopsis activates components of the innate immune response

Huffaker, Alisa; Pearce, Gregory; Ryan, Clarence A.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
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16.32%
Innate immunity is initiated in animals and plants through the recognition of a variety of pathogen-associated molecules that in animals are called pathogen-associated molecular patterns and in plants are called elicitors. Some plant pathogen-derived elicitors have been identified as peptides, but peptide elicitors derived from the plant itself that activate defensive genes against pathogens have not been previously identified. Here, we report the isolation and characterization of a 23-aa peptide from Arabidopsis, called AtPep1, which activates transcription of the defensive gene defensin (PDF1.2) and activates the synthesis of H2O2, both being components of the innate immune response. The peptide is derived from a 92-aa precursor encoded within a small gene that is inducible by wounding, methyl jasmonate, and ethylene. Constitutive expression of the AtPep1 precursor gene PROPEP1 in transgenic Arabidopsis plants causes a constitutive transcription of PDF1.2. When grown in soil, the transgenic plants exhibited an increased root development compared with WT plants and an enhanced resistance toward the root pathogen Pythium irregulare. Six paralogs of PROPEP1 are present in Arabidopsis, and orthologs have been identified in species of several agriculturally important plant families...

The cell surface leucine-rich repeat receptor for AtPep1, an endogenous peptide elicitor in Arabidopsis, is functional in transgenic tobacco cells

Yamaguchi, Yube; Pearce, Gregory; Ryan, Clarence A.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
26.89%
AtPep1 is a 23-aa endogenous peptide elicitor from Arabidopsis leaves that signals the activation of components of the innate immune response against pathogens. Here, we report the isolation of an AtPep1 receptor from the surface of Arabidopsis suspension-cultured cells. An 125I-labeled AtPep1 analog interacted with suspension-cultured Arabidopsis with a Kd of 0.25 nM, and an 125I-labeled azido-Cys-AtPep1 photoaffinity analog specifically labeled a membrane-associated protein of ≈170 kDa. The labeled protein was purified to homogeneity, and its tryptic peptides were identified as gene At1g73080, which encodes a leucine-rich repeat receptor kinase, here called PEPR1. Verification of the binding protein as the receptor for AtPep1 was established by demonstrating the loss of function of microsomal membranes of two SALK insertional mutants and by a gain in function of the alkalinization response to AtPep1 by tobacco suspension-cultured cells expressing the At1g73080 transgene. Synthetic homologs of AtPep1, deduced from the C termini of six known paralogs of PROPEP1, were biologically active and were competitors of the interaction of an AtPep1 radiolabeled analog with the receptor. The data are consistent with a role for PEPR1 as the receptor for AtPep1 to amplify innate immunity in response to pathogen attacks.

Endogenous peptide defense signals in Arabidopsis differentially amplify signaling for the innate immune response

Huffaker, Alisa; Ryan, Clarence A.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
15.96%
AtPep1, a 23-aa peptide encoded by Arabidopsis PROPEP1, a member of a small, six-member gene family, activates expression of the defense gene PDF1.2 (encoding defensin) and its own precursor gene, PROPEP1, through the jasmonate/ethylene signaling pathway, mediated by a cell-surface receptor, PEPR1. Overexpression of two family members, PROPEP1 and PROPEP2, enhances resistance of Arabidopsis plants against the pathogen Pythium irregulare, and PROPEP2 and PROPEP3 are expressed at highly elevated levels in Arabidopsis in response to pathogen infections and to several pathogen-associated molecules (general elicitors). Here, we report that PDF1.2, PR-1 (pathogenesis protein), and PROPEP genes were differentially expressed in the leaves of intact plants sprayed with methyl jasmonate and methyl salicylate and in excised leaves supplied through cut petioles with peptides derived from the C terminus of each of the encoded proteins. The expression of PDF1.2 and PR-1 elicited by the peptides was blocked in mutant plants deficient in the jasmonate/ethylene and salicylate pathways, and in wild-type plants by treatment with diphenylene iodonium chloride, an inhibitor of hydrogen peroxide production. PROPEP1, PROPEP 2, and PROPEP3 genes appear to have roles in a feedback loop that amplifies defense signaling pathways initiated by pathogens.

Rapid Heteromerization and Phosphorylation of Ligand-activated Plant Transmembrane Receptors and Their Associated Kinase BAK1*

Schulze, Birgit; Mentzel, Tobias; Jehle, Anna K.; Mueller, Katharina; Beeler, Seraina; Boller, Thomas; Felix, Georg; Chinchilla, Delphine
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
15.96%
In plants leucine-rich repeat receptor kinases (LRR-RKs) located at the plasma membrane play a pivotal role in the perception of extracellular signals. For two of these LRR-RKs, the brassinosteroid receptor BRI1 and the flagellin receptor FLS2, interaction with the LRR receptor-like kinase BAK1 (BRI1-associated receptor kinase 1) was shown to be required for signal transduction. Here we report that FLS2·BAK1 heteromerization occurs almost instantaneously after perception of the ligand, the flagellin-derived peptide flg22. Flg22 can induce formation of a stable FLS2·BAK1 complex in microsomal membrane preparations in vitro, and the kinase inhibitor K-252a does not prevent complex formation. A kinase dead version of BAK1 associates with FLS2 in a flg22-dependent manner but does not restore responsiveness to flg22 in cells of bak1 plants, demonstrating that kinase activity of BAK1 is essential for FLS2 signaling. Furthermore, using in vivo phospholabeling, we are able to detect de novo phosphorylation of both FLS2 and BAK1 within 15 s of stimulation with flg22. Similarly, brassinolide induces BAK1 phosphorylation within seconds. Other triggers of plant defense, such as bacterial EF-Tu and the endogenous AtPep1 likewise induce rapid formation of heterocomplexes consisting of de novo phosphorylated BAK1 and proteins representing the ligand-specific binding receptors EF-Tu receptor and Pep1 receptor 1...

Perception of the Arabidopsis Danger Signal Peptide 1 Involves the Pattern Recognition Receptor AtPEPR1 and Its Close Homologue AtPEPR2*

Krol, Elzbieta; Mentzel, Tobias; Chinchilla, Delphine; Boller, Thomas; Felix, Georg; Kemmerling, Birgit; Postel, Sandra; Arents, Michael; Jeworutzki, Elena; Al-Rasheid, Khaled A. S.; Becker, Dirk; Hedrich, Rainer
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
16.71%
Plasma membrane-borne pattern recognition receptors, which recognize microbe-associated molecular patterns and endogenous damage-associated molecular patterns, provide the first line of defense in innate immunity. In plants, leucine-rich repeat receptor kinases fulfill this role, as exemplified by FLS2 and EFR, the receptors for the microbe-associated molecular patterns flagellin and elongation factor Tu. Here we examined the perception of the damage-associated molecular pattern peptide 1 (AtPep1), an endogenous peptide of Arabidopsis identified earlier and shown to be perceived by the leucine-rich repeat protein kinase PEPR1. Using seedling growth inhibition, elicitation of an oxidative burst and induction of ethylene biosynthesis, we show that wild type plants and the pepr1 and pepr2 mutants, affected in PEPR1 and in its homologue PEPR2, are sensitive to AtPep1, but that the double mutant pepr1/pepr2 is completely insensitive. As a central body of our study, we provide electrophysiological evidence that at the level of the plasma membrane, AtPep1 triggers a receptor-dependent transient depolarization through activation of plasma membrane anion channels, and that this effect is absent in the double mutant pepr1/pepr2. The double mutant also fails to respond to AtPep2 and AtPep3...

ZmPep1, an Ortholog of Arabidopsis Elicitor Peptide 1, Regulates Maize Innate Immunity and Enhances Disease Resistance1[W][OA]

Huffaker, Alisa; Dafoe, Nicole J.; Schmelz, Eric A.
Fonte: American Society of Plant Biologists Publicador: American Society of Plant Biologists
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
16.32%
ZmPep1 is a bioactive peptide encoded by a previously uncharacterized maize (Zea mays) gene, ZmPROPEP1. ZmPROPEP1 was identified by sequence similarity as an ortholog of the Arabidopsis (Arabidopsis thaliana) AtPROPEP1 gene, which encodes the precursor protein of elicitor peptide 1 (AtPep1). Together with its receptors, AtPEPR1 and AtPEPR2, AtPep1 functions to activate and amplify innate immune responses in Arabidopsis and enhances resistance to both Pythium irregulare and Pseudomonas syringae. Candidate orthologs to the AtPROPEP1 gene have been identified from a variety of crop species; however, prior to this study, activities of the respective peptides encoded by these orthologs were unknown. Expression of the ZmPROPEP1 gene is induced by fungal infection and treatment with jasmonic acid or ZmPep1. ZmPep1 activates de novo synthesis of the hormones jasmonic acid and ethylene and induces the expression of genes encoding the defense proteins endochitinase A, PR-4, PRms, and SerPIN. ZmPep1 also stimulates the expression of Benzoxazineless1, a gene required for the biosynthesis of benzoxazinoid defenses, and the accumulation of 2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one glucoside in leaves. To ascertain whether ZmPep1-induced defenses affect resistance...

Cyclic nucleotide gated channel gene family in tomato: genome-wide identification and functional analyses in disease resistance

Saand, Mumtaz A.; Xu, You-Ping; Li, Wen; Wang, Ji-Peng; Cai, Xin-Zhong
Fonte: Frontiers Media S.A. Publicador: Frontiers Media S.A.
Tipo: Artigo de Revista Científica
Publicado em 05/05/2015 EN
Relevância na Pesquisa
15.96%
The cyclic nucleotide gated channel (CNGC) is suggested to be one of the important calcium conducting channels. Nevertheless, genome-wide identification and systemic functional analysis of CNGC gene family in crop plant species have not yet been conducted. In this study, we performed genome-wide identification of CNGC gene family in the economically important crop tomato (Solanum lycopersicum L.) and analyzed function of the group IVb SlCNGC genes in disease resistance. Eighteen CNGC genes were identified in tomato genome, and four CNGC loci that were misannotated at database were corrected by cloning and sequencing. Detailed bioinformatics analyses on gene structure, domain composition and phylogenetic relationship of the SlCNGC gene family were conducted and the group-specific feature was revealed. Comprehensive expression analyses demonstrated that SlCNGC genes were highly, widely but differently responsive to diverse stimuli. Pharmacological assays showed that the putative CNGC activators cGMP and cAMP enhanced resistance against Sclerotinia sclerotiorum. Silencing of group IVb SlCNGC genes significantly enhanced resistance to fungal pathogens Pythium aphanidermatum and S. sclerotiorum, strongly reduced resistance to viral pathogen Tobacco rattle virus...

Analysis on the BAK1-interacting RLKs BIR2 and BIR3, two proteins that differentially regulate BAK1-dependent pathways

Imkampe, Julia
Fonte: Universität Tübingen Publicador: Universität Tübingen
Tipo: Dissertation; info:eu-repo/semantics/doctoralThesis
EN
Relevância na Pesquisa
15.96%
BAK1 is a multifunctional co-receptor that positively regulates multiple plant signaling pathways by interacting with the corresponding ligand-binding receptors, e.g. BRI1, FLS2, EFR, PEPR1 and PEPR2. Moreover, BAK1 plays a role in cell death control. In order to investigate how BAK1 is regulated we purified in vivo BAK1 complexes and identified two new BAK1 interactors, the BAK1 interacting RLKs 2 and 3 (BIR2 and BIR3). Functional analysis of bir2 mutants showed that they are hyperresponsive to MAMP treatment, show cell death spreading after pathogen infection but are not impaired in BL signaling. In this study the molecular mechanism how BIR2 regulates MAMP responses was further investigated. With Co-IP analysis in Arabidopsis wildtype plants it was shown that treatment with flg22 leads to a partial release of BAK1 from BIR2. Treatment with a cocktail of flg22, elf18, BL and Atpep1 leads to increased release of BAK1 from BIR2 compared to single treatments, indicating that BAK1 exists in preformed complexes with different ligand-binding receptors. Functional analyses of 35S-BIR3 plants have shown that BIR3 is a negative regulator of BL responses, MAMP responses and cell death. In this study the role of BIR3 in BL responses was further investigated and it could be shown that BIR3 not only interacts with BAK1 but also shows direct interaction with the ligand-binding receptor BRI1. The involvement of BIR3 in MAMP responses was confirmed with different functional assays. On the molecular level it was shown that BIR3 interacts with BAK1 and FLS2 and regulates BAK1-FLS2 complex formation. bak1 bir3 double mutants show a strong growth phenotype with spontaneous lesion formation and strong cell death spreading after A. brassicicola infections...