The RING E3 Ligase KEEP ON GOING Modulates JASMONATE ZIM-DOMAIN12 Stability1[OPEN]

An E3 ubiquitin ligase involved in abscisic acid signaling modulates the stability of a central jasmonate signaling component. Jasmonate (JA) signaling in plants is mediated by the JASMONATE ZIM-DOMAIN (JAZ) proteins that repress the activity of several transcription factors regulating JA-inducible gene expression. The hormone JA-isoleucine triggers the interaction of JAZ repressor proteins with the F-box protein CORONATINE INSENSITIVE1 (COI1), part of an S-phase kinase-associated protein1/Cullin1/F-box protein COI1 (SCFCOI1) E3 ubiquitin ligase complex, and their degradation by the 26S proteasome. In Arabidopsis (Arabidopsis thaliana), the JAZ family consists of 13 members. The level of redundancy or specificity among these members is currently not well understood. Here, we characterized JAZ12, encoded by a highly expressed JAZ gene. JAZ12 interacted with the transcription factors MYC2, MYC3, and MYC4 in vivo and repressed MYC2 activity. Using tandem affinity purification, we found JAZ12 to interact with SCFCOI1 components, matching with observed in vivo ubiquitination and with rapid degradation after treatment with JA. In contrast to the other JAZ proteins, JAZ12 also interacted directly with the E3 RING ligase KEEP ON GOING (KEG), a known repressor of the ABSCISIC ACID INSENSITIVE5 transcription factor in abscisic acid signaling. To study the functional role of this interaction, we circumvented the lethality of keg loss-of-function mutants by silencing KEG using an artificial microRNA approach. Abscisic acid treatment promoted JAZ12 degradation, and KEG knockdown led to a decrease in JAZ12 protein levels. Correspondingly, KEG overexpression was capable of partially inhibiting COI1-mediated JAZ12 degradation. Our results provide additional evidence for KEG as an important factor in plant hormone signaling and a positive regulator of JAZ12 stability.

[1]  Anna Wawrzyńska,et al.  Powdery Mildew Resistance Conferred by Loss of the ENHANCED DISEASE RESISTANCE1 Protein Kinase Is Suppressed by a Missense Mutation in KEEP ON GOING, a Regulator of Abscisic Acid Signaling1[W][OA] , 2008, Plant Physiology.

[2]  K. Nakamura,et al.  Sugar-Inducible Expression of a Gene for [beta]-Amylase in Arabidopsis thaliana , 1995, Plant physiology.

[3]  A. J. Koo,et al.  Temporal Dynamics of Growth and Photosynthesis Suppression in Response to Jasmonate Signaling1[W][OPEN] , 2014, Plant Physiology.

[4]  Hong Ma,et al.  The SCF(COI1) ubiquitin-ligase complexes are required for jasmonate response in Arabidopsis. , 2002, The Plant cell.

[5]  Jonathan Legrand,et al.  A fluorescent hormone biosensor reveals the dynamics of jasmonate signalling in plants , 2015, Nature Communications.

[6]  R. Vierstra,et al.  KEEP ON GOING, a RING E3 Ligase Essential for Arabidopsis Growth and Development, Is Involved in Abscisic Acid Signaling[W] , 2006, The Plant Cell Online.

[7]  G. Howe,et al.  Repression of jasmonate signaling by a non-TIFY JAZ protein in Arabidopsis. , 2015, The Plant journal : for cell and molecular biology.

[8]  J. Franco-Zorrilla,et al.  The Arabidopsis bHLH Transcription Factors MYC3 and MYC4 Are Targets of JAZ Repressors and Act Additively with MYC2 in the Activation of Jasmonate Responses[C][W] , 2011, Plant Cell.

[9]  Yangnan Gu,et al.  The KEEP ON GOING Protein of Arabidopsis Recruits the ENHANCED DISEASE RESISTANCE1 Protein to Trans-Golgi Network/Early Endosome Vesicles1[W][OA] , 2011, Plant Physiology.

[10]  J. Manners,et al.  JAZ repressors and the orchestration of phytohormone crosstalk. , 2012, Trends in plant science.

[11]  Hongwei Guo,et al.  The bHLH Subgroup IIId Factors Negatively Regulate Jasmonate-Mediated Plant Defense and Development , 2013, PLoS genetics.

[12]  S. Davis,et al.  TIME FOR COFFEE Represses Accumulation of the MYC2 Transcription Factor to Provide Time-of-Day Regulation of Jasmonate Signaling in Arabidopsis[C][W][OA] , 2012, Plant Cell.

[13]  D. Baulcombe,et al.  An enhanced transient expression system in plants based on suppression of gene silencing by the p19 protein of tomato bushy stunt virus. , 2003, The Plant journal : for cell and molecular biology.

[14]  D. Inzé,et al.  Mapping methyl jasmonate-mediated transcriptional reprogramming of metabolism and cell cycle progression in cultured Arabidopsis cells , 2008, Proceedings of the National Academy of Sciences.

[15]  R. Solano,et al.  JASMONATE-INSENSITIVE1 Encodes a MYC Transcription Factor Essential to Discriminate between Different Jasmonate-Regulated Defense Responses in Arabidopsis , 2004, The Plant Cell Online.

[16]  Kazuo Shinozaki,et al.  Arabidopsis AtMYC2 (bHLH) and AtMYB2 (MYB) Function as Transcriptional Activators in Abscisic Acid Signaling Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.006130. , 2003, The Plant Cell Online.

[17]  A. Hills,et al.  EZ-Rhizo: integrated software for the fast and accurate measurement of root system architecture. , 2009, The Plant journal : for cell and molecular biology.

[18]  D. Inzé,et al.  Systematic Localization of the Arabidopsis Core Cell Cycle Proteins Reveals Novel Cell Division Complexes1[W][OA] , 2009, Plant Physiology.

[19]  Bryan C Thines,et al.  JAZ repressor proteins are targets of the SCFCOI1 complex during jasmonate signalling , 2007, Nature.

[20]  Steven P. Gygi,et al.  Weighing in on ubiquitin: the expanding role of mass-spectrometry-based proteomics , 2005, Nature Cell Biology.

[21]  G. Howe,et al.  A critical role of two positively charged amino acids in the Jas motif of Arabidopsis JAZ proteins in mediating coronatine- and jasmonoyl isoleucine-dependent interactions with the COI1 F-box protein. , 2008, The Plant journal : for cell and molecular biology.

[22]  D. Inzé,et al.  NINJA connects the co-repressor TOPLESS to jasmonate signalling , 2010, Nature.

[23]  A. Goossens,et al.  The JAZ Proteins: A Crucial Interface in the Jasmonate Signaling Cascade , 2011, Plant Cell.

[24]  Wei Li,et al.  Derepression of ethylene-stabilized transcription factors (EIN3/EIL1) mediates jasmonate and ethylene signaling synergy in Arabidopsis , 2011, Proceedings of the National Academy of Sciences.

[25]  Marta Godoy,et al.  ABA Is an Essential Signal for Plant Resistance to Pathogens Affecting JA Biosynthesis and the Activation of Defenses in Arabidopsis[W] , 2007, The Plant Cell Online.

[26]  S. Stolz,et al.  Role of NINJA in root jasmonate signaling , 2013, Proceedings of the National Academy of Sciences.

[27]  A. Goossens,et al.  Change of a conserved amino acid in the MYC2 and MYC3 transcription factors leads to release of JAZ repression and increased activity. , 2015, The New phytologist.

[28]  S. Mita,et al.  Sugar-lnducible Expression of a Gene for P-Amylase in Arabidopsis thaliana ' , 2002 .

[29]  Hoo Sun Chung,et al.  Negative Feedback Control of Jasmonate Signaling by an Alternative Splice Variant of JAZ101[C][W][OA] , 2013, Plant Physiology.

[30]  G. Howe,et al.  Transcription factor-dependent nuclear localization of a transcriptional repressor in jasmonate hormone signaling , 2012, Proceedings of the National Academy of Sciences.

[31]  Detlef Weigel,et al.  Highly Specific Gene Silencing by Artificial MicroRNAs in Arabidopsis[W][OA] , 2006, The Plant Cell Online.

[32]  J. Franco-Zorrilla,et al.  bHLH003, bHLH013 and bHLH017 Are New Targets of JAZ Repressors Negatively Regulating JA Responses , 2014, PloS one.

[33]  P. Hilson,et al.  Modular cloning in plant cells. , 2005, Trends in plant science.

[34]  C. Pieterse,et al.  The Non-JAZ TIFY Protein TIFY8 from Arabidopsis thaliana Is a Transcriptional Repressor , 2014, PloS one.

[35]  A. J. Koo,et al.  A bHLH-Type Transcription Factor, ABA-INDUCIBLE BHLH-TYPE TRANSCRIPTION FACTOR/JA-ASSOCIATED MYC2-LIKE1, Acts as a Repressor to Negatively Regulate Jasmonate Signaling in Arabidopsis[C][W] , 2013, Plant Cell.

[36]  Serban Nacu,et al.  Fast and SNP-tolerant detection of complex variants and splicing in short reads , 2010, Bioinform..

[37]  R. Solano,et al.  Repression of Jasmonate-Dependent Defenses by Shade Involves Differential Regulation of Protein Stability of MYC Transcription Factors and Their JAZ Repressors in Arabidopsis[C][W] , 2014, Plant Cell.

[38]  A. Goossens,et al.  Transient expression assays in tobacco protoplasts. , 2013, Methods in molecular biology.

[39]  Yangnan Gu,et al.  The KEEP ON GOING Protein of Arabidopsis Regulates Intracellular Protein Trafficking and Is Degraded during Fungal Infection[C][W][OA] , 2012, Plant Cell.

[40]  Lennart Martens,et al.  An improved toolbox to unravel the plant cellular machinery by tandem affinity purification of Arabidopsis protein complexes , 2014, Nature Protocols.

[41]  D. Inzé,et al.  An abscisic acid-sensitive checkpoint in lateral root development of Arabidopsis. , 2003, The Plant journal : for cell and molecular biology.

[42]  S. Stone,et al.  Cytoplasmic Degradation of the Arabidopsis Transcription Factor ABSCISIC ACID INSENSITIVE 5 Is Mediated by the RING-type E3 Ligase KEEP ON GOING* , 2013, The Journal of Biological Chemistry.

[43]  Cynthia D. Nezames,et al.  ABD1 Is an Arabidopsis DCAF Substrate Receptor for CUL4-DDB1–Based E3 Ligases That Acts as a Negative Regulator of Abscisic Acid Signaling[W] , 2014, Plant Cell.

[44]  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.

[45]  Lloyd M. Smith,et al.  Tandem affinity purification and mass spectrometric analysis of ubiquitylated proteins in Arabidopsis. , 2009, The Plant journal : for cell and molecular biology.

[46]  Fang Wang,et al.  Jasmonate Regulates the INDUCER OF CBF EXPRESSION–C-REPEAT BINDING FACTOR/DRE BINDING FACTOR1 Cascade and Freezing Tolerance in Arabidopsis[W] , 2013, Plant Cell.

[47]  Hoo Sun Chung,et al.  A Critical Role for the TIFY Motif in Repression of Jasmonate Signaling by a Stabilized Splice Variant of the JASMONATE ZIM-Domain Protein JAZ10 in Arabidopsis[C][W] , 2009, The Plant Cell Online.

[48]  Q. Xie,et al.  The Arabidopsis F-Box Protein CORONATINE INSENSITIVE1 Is Stabilized by SCF COI1 and Degraded via the 26S , 2013 .

[49]  I. Hara-Nishimura,et al.  A rapid and non-destructive screenable marker, FAST, for identifying transformed seeds of Arabidopsis thaliana. , 2010, The Plant journal : for cell and molecular biology.

[50]  D. Inzé,et al.  Expression of the Arabidopsis jasmonate signalling repressor JAZ1/TIFY10A is stimulated by auxin , 2009, EMBO reports.

[51]  W. Terzaghi,et al.  DWA1 and DWA2, Two Arabidopsis DWD Protein Components of CUL4-Based E3 Ligases, Act Together as Negative Regulators in ABA Signal Transduction[C][W] , 2010, Plant Cell.

[52]  M. Hamberg,et al.  (+)-7-iso-Jasmonoyl-L-isoleucine is the endogenous bioactive jasmonate. , 2009, Nature chemical biology.

[53]  M. Van Montagu,et al.  Jasmonate signaling involves the abscisic acid receptor PYL4 to regulate metabolic reprogramming in Arabidopsis and tobacco , 2011, Proceedings of the National Academy of Sciences.

[54]  Q. Xie,et al.  The Arabidopsis F-Box Protein CORONATINE INSENSITIVE1 Is Stabilized by SCFCOI1 and Degraded via the 26S Proteasome Pathway[C][W] , 2013, Plant Cell.

[55]  A. Schofield,et al.  Arabidopsis CIPK26 interacts with KEG, components of the ABA signalling network and is degraded by the ubiquitin–proteasome system , 2013, Journal of experimental botany.

[56]  J. Callis,et al.  ABA and the ubiquitin E3 ligase KEEP ON GOING affect proteolysis of the Arabidopsis thaliana transcription factors ABF1 and ABF3 , 2013, The Plant journal : for cell and molecular biology.

[57]  Paul Theodor Pyl,et al.  HTSeq—a Python framework to work with high-throughput sequencing data , 2014, bioRxiv.

[58]  H. Ohta,et al.  Basic Helix-Loop-Helix Transcription Factors JASMONATE-ASSOCIATED MYC2-LIKE1 (JAM1), JAM2, and JAM3 Are Negative Regulators of Jasmonate Responses in Arabidopsis1[W][OPEN] , 2013, Plant Physiology.

[59]  M. Pagni,et al.  A Downstream Mediator in the Growth Repression Limb of the Jasmonate Pathway[W][OA] , 2007, The Plant Cell Online.

[60]  Hoo Sun Chung,et al.  Comprehensive Protein-Based Artificial MicroRNA Screens for Effective Gene Silencing in Plants[W] , 2013, Plant Cell.

[61]  Paul R. Ebert,et al.  Antagonistic Interaction between Abscisic Acid and Jasmonate-Ethylene Signaling Pathways Modulates Defense Gene Expression and Disease Resistance in Arabidopsis , 2004, The Plant Cell Online.

[62]  Dirk Inzé,et al.  Boosting tandem affinity purification of plant protein complexes. , 2008, Trends in plant science.

[63]  Anthony L. Schilmiller,et al.  COI1 is a critical component of a receptor for jasmonate and the bacterial virulence factor coronatine , 2008, Proceedings of the National Academy of Sciences.

[64]  A. Goossens,et al.  Yeast two-hybrid analysis of jasmonate signaling proteins. , 2013, Methods in molecular biology.

[65]  P. Figueroa,et al.  Characterization of JAZ-interacting bHLH transcription factors that regulate jasmonate responses in Arabidopsis , 2011, Journal of experimental botany.

[66]  J. Rizo,et al.  Jasmonate perception by inositol phosphate-potentiated COI1-JAZ co-receptor , 2010, Nature.

[67]  J. Micol,et al.  The JAZ family of repressors is the missing link in jasmonate signalling , 2007, Nature.

[68]  G. Howe,et al.  JAZ8 Lacks a Canonical Degron and Has an EAR Motif That Mediates Transcriptional Repression of Jasmonate Responses in Arabidopsis[C][W] , 2012, Plant Cell.

[69]  T. Lynch,et al.  A small plant-specific protein family of ABI five binding proteins (AFPs) regulates stress response in germinating Arabidopsis seeds and seedlings , 2008, Plant Molecular Biology.

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

[71]  S. Stone,et al.  Abscisic Acid Increases Arabidopsis ABI5 Transcription Factor Levels by Promoting KEG E3 Ligase Self-Ubiquitination and Proteasomal Degradation[W][OA] , 2010, Plant Cell.

[72]  M. Dong,et al.  Phosphorylation-Coupled Proteolysis of the Transcription Factor MYC2 Is Important for Jasmonate-Signaled Plant Immunity , 2013, PLoS genetics.