Ethylene-responsive element-binding factor 5, ERF5, is involved in chitin-induced innate immunity response.

Our recent work demonstrated that chitin treatment modulated the expression of 118 transcription factor (TF) genes in Arabidopsis. To investigate the potential roles of these TF in chitin signaling and plant defense, we initiated an interaction study among these TF proteins, as well as two chitin-activated mitogen-activated protein kinases (MPK3 and MPK6), using a yeast two-hybrid system. This study revealed interactions among the following proteins: three ethylene-responsive element-binding factors (ERF), five WRKY transcription factors, one scarecrow-like (SCL), and the two MPK, in addition to many other interactions, reflecting a complex TF interaction network. Most of these interactions were subsequently validated by other methods, such as pull-down and in planta bimolecular fluorescence complementation assays. The key node ERF5 was shown to interact with multiple proteins in the network, such as ERF6, ERF8, and SCL13, as well as MPK3 and MPK6. Interestingly, ERF5 appeared to negatively regulate chitin signaling and plant defense against the fungal pathogen Alternaria brassicicola and positively regulate salicylic acid signaling and plant defense against the bacterial pathogen Pseudomonas syringae pv. tomato DC3000. Therefore, ERF5 may play an important role in plant innate immunity, likely through coordinating chitin and other defense pathways in plants in response to different pathogens.

[1]  G. Stacey,et al.  Activation of a mitogen-activated protein kinase pathway in Arabidopsis by chitin. , 2004, Molecular plant pathology.

[2]  T. Eulgem.  Dissecting the WRKY Web of Plant Defense Regulators , 2006, PLoS pathogens.

[3]  K. Hiratsu,et al.  Dominant repression of target genes by chimeric repressors that include the EAR motif, a repression domain, in Arabidopsis. , 2003, The Plant journal : for cell and molecular biology.

[4]  K. Hiratsu,et al.  Repression Domains of Class II ERF Transcriptional Repressors Share an Essential Motif for Active Repression , 2001, The Plant Cell Online.

[5]  T. Fujimura,et al.  Genome-Wide Analysis of the ERF Gene Family in Arabidopsis and Rice[W] , 2006, Plant Physiology.

[6]  Martin J. Mueller,et al.  NPR1 Modulates Cross-Talk between Salicylate- and Jasmonate-Dependent Defense Pathways through a Novel Function in the Cytosol Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.009159. , 2003, The Plant Cell Online.

[7]  M. Mitsutomi,et al.  Direct Binding of a Plant LysM Receptor-like Kinase, LysM RLK1/CERK1, to Chitin in Vitro* , 2009, The Journal of Biological Chemistry.

[8]  M. Ohme-Takagi,et al.  Arabidopsis Ethylene-Responsive Element Binding Factors Act as Transcriptional Activators or Repressors of GCC Box–Mediated Gene Expression , 2000, Plant Cell.

[9]  P. Agarwal,et al.  Role of DREB transcription factors in abiotic and biotic stress tolerance in plants , 2006, Plant Cell Reports.

[10]  J. Thorner,et al.  Function and regulation in MAPK signaling pathways: lessons learned from the yeast Saccharomyces cerevisiae. , 2007, Biochimica et biophysica acta.

[11]  J. Chory,et al.  Weaving the complex web of signal transduction. , 2001, Plant physiology.

[12]  I. Somssich,et al.  The Role of WRKY Transcription Factors in Plant Immunity[W] , 2009, Plant Physiology.

[13]  G. Stacey,et al.  A LysM Receptor-Like Kinase Plays a Critical Role in Chitin Signaling and Fungal Resistance in Arabidopsis[W][OA] , 2008, The Plant Cell Online.

[14]  I. Somssich,et al.  A new member of the Arabidopsis WRKY transcription factor family, AtWRKY6, is associated with both senescence- and defence-related processes. , 2001, The Plant journal : for cell and molecular biology.

[15]  M. Schmid,et al.  Genome-Wide Insertional Mutagenesis of Arabidopsis thaliana , 2003, Science.

[16]  Imre E Somssich,et al.  Targets of AtWRKY6 regulation during plant senescence and pathogen defense. , 2002, Genes & development.

[17]  A. Gavin,et al.  Interaction networks for systems biology , 2008, FEBS letters.

[18]  Paul J. Choi,et al.  Bimolecular fluorescence complementation (BiFC) analysis as a probe of protein interactions in living cells. , 2008, Annual review of biophysics.

[19]  Alexandra To,et al.  A Network of Local and Redundant Gene Regulation Governs Arabidopsis Seed Maturation , 2006, The Plant Cell Online.

[20]  I. Somssich,et al.  Expression of AtWRKY33 encoding a pathogen- or PAMP-responsive WRKY transcription factor is regulated by a composite DNA motif containing W box elements. , 2007, Molecular plant-microbe interactions : MPMI.

[21]  G. Stacey,et al.  Chitin signaling and plant disease resistance , 2008, Plant signaling & behavior.

[22]  W. Broekaert,et al.  The role of ethylene in host-pathogen interactions. , 2006, Annual review of phytopathology.

[23]  T. Mengiste,et al.  Arabidopsis WRKY33 transcription factor is required for resistance to necrotrophic fungal pathogens. , 2006, The Plant journal : for cell and molecular biology.

[24]  G. Stacey,et al.  Loss-of-Function Mutations in Chitin Responsive Genes Show Increased Susceptibility to the Powdery Mildew Pathogen Erysiphe cichoracearum1[w] , 2005, Plant Physiology.

[25]  B. Kunkel,et al.  Cross talk between signaling pathways in pathogen defense. , 2002, Current opinion in plant biology.

[26]  J. Uhrig,et al.  Protein interaction networks in plants , 2006, Planta.

[27]  Tetsuro Mimura,et al.  Transcription switches for protoxylem and metaxylem vessel formation. , 2005, Genes & development.

[28]  Jing Li,et al.  The WRKY70 Transcription Factor: A Node of Convergence for Jasmonate-Mediated and Salicylate-Mediated Signals in Plant Defense On-line version contains Web-only data. , 2004, The Plant Cell Online.

[29]  Kengo Kinoshita,et al.  ATTED-II provides coexpressed gene networks for Arabidopsis , 2008, Nucleic Acids Res..

[30]  Moisés Santillán,et al.  Comparative analysis of the transcription-factor gene regulatory networks of E. coli and S. cerevisiae , 2008, BMC Systems Biology.

[31]  A. Vainstein,et al.  Subcellular localization of interacting proteins by bimolecular fluorescence complementation in planta. , 2006, Journal of molecular biology.

[32]  Pradeep Kachroo,et al.  Salicylic acid-, jasmonic acid- and ethylene-mediated regulation of plant defense signaling. , 2007, Genetic engineering.

[33]  Yoko Nishizawa,et al.  Plant cells recognize chitin fragments for defense signaling through a plasma membrane receptor. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[34]  F. Ausubel,et al.  MAP kinase signalling cascade in Arabidopsis innate immunity , 2002, Nature.

[35]  H. Hirt,et al.  The MAP kinase substrate MKS1 is a regulator of plant defense responses. , 2005, The EMBO journal.

[36]  Christian A. Grove,et al.  Transcription factor functionality and transcription regulatory networks. , 2008, Molecular bioSystems.

[37]  G. Stacey,et al.  Identification of 118 Arabidopsis transcription factor and 30 ubiquitin-ligase genes responding to chitin, a plant-defense elicitor. , 2007, Molecular plant-microbe interactions : MPMI.

[38]  B. Thomma,et al.  Separate jasmonate-dependent and salicylate-dependent defense-response pathways in Arabidopsis are essential for resistance to distinct microbial pathogens. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[39]  T. Kerppola.  Visualization of molecular interactions using bimolecular fluorescence complementation analysis: characteristics of protein fragment complementation. , 2009, Chemical Society reviews.

[40]  Zhixiang Chen,et al.  Potentiation of Developmentally Regulated Plant Defense Response by AtWRKY18, a Pathogen-Induced Arabidopsis Transcription Factor1 , 2002, Plant Physiology.

[41]  J. Ecker,et al.  Exploiting the triple response of Arabidopsis to identify ethylene-related mutants. , 1990, The Plant cell.

[42]  T. Boller,et al.  Perception of the Bacterial PAMP EF-Tu by the Receptor EFR Restricts Agrobacterium-Mediated Transformation , 2006, Cell.

[43]  G. Stacey,et al.  Characterization of early, chitin-induced gene expression in Arabidopsis. , 2002, Molecular plant-microbe interactions : MPMI.

[44]  Rachel B. Brem,et al.  Integrating large-scale functional genomic data to dissect the complexity of yeast regulatory networks , 2008, Nature Genetics.

[45]  Synan F. AbuQamar,et al.  The Membrane-Anchored BOTRYTIS-INDUCED KINASE1 Plays Distinct Roles in Arabidopsis Resistance to Necrotrophic and Biotrophic Pathogens[W] , 2005, The Plant Cell Online.

[46]  Caren Chang,et al.  Ethylene signaling: new levels of complexity and regulation. , 2008, Current opinion in plant biology.

[47]  Yong Li,et al.  An Arabidopsis thaliana T-DNA mutagenized population (GABI-Kat) for flanking sequence tag-based reverse genetics , 2003, Plant Molecular Biology.

[48]  Y. Narusaka,et al.  CERK1, a LysM receptor kinase, is essential for chitin elicitor signaling in Arabidopsis , 2007, Proceedings of the National Academy of Sciences.

[49]  T. Eulgem,et al.  Networks of WRKY transcription factors in defense signaling. , 2007, Current opinion in plant biology.

[50]  Veena,et al.  IMPa-4, an Arabidopsis Importin α Isoform, Is Preferentially Involved in Agrobacterium-Mediated Plant Transformation[W] , 2008, The Plant Cell Online.

[51]  S. Clough,et al.  Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. , 1998, The Plant journal : for cell and molecular biology.

[52]  C. Pieterse,et al.  The AP2/ERF Domain Transcription Factor ORA59 Integrates Jasmonic Acid and Ethylene Signals in Plant Defense1[W] , 2008, Plant Physiology.

[53]  David L Robertson,et al.  Choose your partners: dimerization in eukaryotic transcription factors. , 2008, Trends in biochemical sciences.

[54]  T. Eulgem,et al.  WRKY72-type transcription factors contribute to basal immunity in tomato and Arabidopsis as well as gene-for-gene resistance mediated by the tomato R gene Mi-1. , 2010, The Plant journal : for cell and molecular biology.

[55]  S. H. Kim,et al.  SRFR1, a suppressor of effector-triggered immunity, encodes a conserved tetratricopeptide repeat protein with similarity to transcriptional repressors. , 2009, The Plant journal : for cell and molecular biology.

[56]  R. Pearson,et al.  Protein kinase phosphorylation site sequences and consensus specificity motifs: tabulations. , 1991, Methods in enzymology.

[57]  R. Mittler,et al.  The zinc finger network of plants , 2008, Cellular and Molecular Life Sciences.

[58]  Xiangzong Meng,et al.  Phosphorylation of a WRKY Transcription Factor by Two Pathogen-Responsive MAPKs Drives Phytoalexin Biosynthesis in Arabidopsis[C][W] , 2011, Plant Cell.

[59]  Kengo Kinoshita,et al.  ATTED-II: a database of co-expressed genes and cis elements for identifying co-regulated gene groups in Arabidopsis , 2006, Nucleic Acids Res..

[60]  E. T. Palva,et al.  WRKY70 modulates the selection of signaling pathways in plant defense. , 2006, The Plant journal : for cell and molecular biology.

[61]  D. Klessig,et al.  MAPK cascades in plant defense signaling. , 2001, Trends in plant science.

[62]  Sorina C. Popescu,et al.  MAPK target networks in Arabidopsis thaliana revealed using functional protein microarrays. , 2009, Genes & development.

[63]  B. Fan,et al.  Physical and Functional Interactions between Pathogen-Induced Arabidopsis WRKY 18 , WRKY 40 , and WRKY 60 Transcription Factors , 2006 .

[64]  F. Ausubel,et al.  Five components of the ethylene-response pathway identified in a screen for weak ethylene-insensitive mutants in Arabidopsis , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[65]  L. C. van Loon,et al.  Ethylene as a modulator of disease resistance in plants. , 2006, Trends in plant science.

[66]  Jane Glazebrook,et al.  Arabidopsis MAP kinase 4 regulates gene expression through transcription factor release in the nucleus , 2008, The EMBO journal.

[67]  M. Vos,et al.  The AP 2 / ERF Domain Transcription Factor ORA 59 Integrates Jasmonic Acid and Ethylene Signals in Plant Defense 1 [ W ] , 2008 .

[68]  Chunhong Chen,et al.  Physical and Functional Interactions between Pathogen-Induced Arabidopsis WRKY18, WRKY40, and WRKY60 Transcription Factors[W] , 2006, The Plant Cell Online.

[69]  E. Minami,et al.  Oligosaccharide signalling for defence responses in plant , 2001 .

[70]  G. Martin,et al.  Role of mitogen-activated protein kinases in plant immunity. , 2005, Current opinion in plant biology.

[71]  U. Grossniklaus,et al.  A Gateway Cloning Vector Set for High-Throughput Functional Analysis of Genes in Planta[w] , 2003, Plant Physiology.

[72]  N. Amornsiripanitch,et al.  A Genomic Approach to Identify Regulatory Nodes in the Transcriptional Network of Systemic Acquired Resistance in Plants , 2006, PLoS pathogens.

[73]  J. Mundy,et al.  Mitogen-activated protein kinase signaling in plants. , 2010, Annual review of plant biology.