Sensing of viral RNA in plants via a DICER-LIKE Ribonuclease

Sensors of intracellular double-stranded RNA are central components of metazoan innate antiviral immunity, but such sensors have not been identified in plants. RNA interference (RNAi) constitutes a potent plant antiviral defense mechanism that relies on conversion of viral RNA into small interfering RNAs by two DICER-LIKE (DCL) ribonucleases, DCL4 and DCL2. Here, we show that while plant DCL4 is dedicated to RNAi, cytoplasmic dicing by DCL2 also triggers RNAi-independent defense gene expression via at least two intracellular nucleotide-binding domain/leucine-rich repeat (NLR) immune receptors. Combined DCL4/NLR inactivation abrogates basal resistance to a positive strand RNA virus. Our results redefine the basis of plant antiviral immunity, including autoimmunity as an explanation for DCL2-dependent growth arrest in dcl and RNA decay mutants in several plant species. One sentence summary The plant immune system uses Dicer-like ribonucleases for both antiviral RNA interference and double-stranded RNA sensing.

[1]  Beixin Mo,et al.  Uridylation and the SKI complex orchestrate the Calvin cycle of photosynthesis through RNA surveillance of TKL1 in Arabidopsis , 2022, Proceedings of the National Academy of Sciences of the United States of America.

[2]  Gil Amitai,et al.  Viruses inhibit TIR gcADPR signaling to overcome bacterial defense , 2022, bioRxiv.

[3]  J. Chai,et al.  Identification and receptor mechanism of TIR-catalyzed small molecules in plant immunity , 2022, bioRxiv.

[4]  P. Schulze-Lefert,et al.  A wheat resistosome defines common principles of immune receptor channels , 2022, bioRxiv.

[5]  R. Panstruga,et al.  Plant autoimmunity—fresh insights into an old phenomenon , 2021, Plant physiology.

[6]  K. Harter,et al.  The EDS1–PAD4–ADR1 node mediates Arabidopsis pattern-triggered immunity , 2021, Nature.

[7]  C. Ritzenthaler,et al.  Immunocapture of dsRNA-bound proteins provides insight into Tobacco rattle virus replication complexes and reveals Arabidopsis DRB2 to be a wide-spectrum antiviral effector , 2021, The Plant cell.

[8]  P. J. Kranzusch,et al.  cGAS-like receptors sense RNA and control 3′2′-cGAMP signalling in Drosophila , 2021, Nature.

[9]  A. Pichlmair,et al.  Two cGAS-like receptors induce antiviral immunity in Drosophila , 2021, Nature.

[10]  Z. Pei,et al.  Plant “helper” immune receptors are Ca2+-permeable nonselective cation channels , 2021, Science.

[11]  Jian-Min Zhou,et al.  The ZAR1 resistosome is a calcium-permeable channel triggering plant immune signaling , 2021, Cell.

[12]  P. Schulze-Lefert,et al.  Direct pathogen-induced assembly of an NLR immune receptor complex to form a holoenzyme , 2020, Science.

[13]  Xiaoqiu Wu,et al.  Structure and function analysis of a CC-NBS-LRR protein AT1G12290. , 2020, Biochemical and biophysical research communications.

[14]  A. Pichlmair,et al.  Human NLRP1 is a sensor for double-stranded RNA , 2020, Science.

[15]  Jonathan D. G. Jones,et al.  Two unequally redundant "helper" immune receptor families mediate Arabidopsis thaliana intracellular "sensor" immune receptor functions , 2020, PLoS biology.

[16]  E. Nogales,et al.  Structure of the activated ROQ1 resistosome directly recognizing the pathogen effector XopQ , 2020, Science.

[17]  J. Rehwinkel,et al.  RIG-I-like receptors: their regulation and roles in RNA sensing , 2020, Nature Reviews Immunology.

[18]  E. Troemel,et al.  The Caenorhabditis elegans RIG-I Homolog DRH-1 Mediates the Intracellular Pathogen Response upon Viral Infection , 2019, Journal of Virology.

[19]  Bostjan Kobe,et al.  NAD+ cleavage activity by animal and plant TIR domains in cell death pathways , 2019, Science.

[20]  Eui-Hwan Chung,et al.  TIR domains of plant immune receptors are NAD+-cleaving enzymes that promote cell death , 2019, Science.

[21]  D. Muruve,et al.  NOD-like receptors and inflammasomes: A review of their canonical and non-canonical signaling pathways. , 2019, Archives of biochemistry and biophysics.

[22]  D. Ren,et al.  Two Arabidopsis Receptor-like Cytoplasmic Kinases SZE1 and SZE2 Associate with the ZAR1-ZED1 Complex and Are Required for Effector-Triggered Immunity. , 2019, Molecular plant.

[23]  J. Vilo,et al.  g:Profiler: a web server for functional enrichment analysis and conversions of gene lists (2019 update) , 2019, Nucleic Acids Res..

[24]  Y. Qi,et al.  Ligand-triggered allosteric ADP release primes a plant NLR complex , 2019, Science.

[25]  Y. Qi,et al.  Reconstitution and structure of a plant NLR resistosome conferring immunity , 2019, Science.

[26]  Li Feng,et al.  A Phytophthora Effector Suppresses Trans-Kingdom RNAi to Promote Disease Susceptibility. , 2019, Cell host & microbe.

[27]  M. Bergdoll,et al.  Characterization of DCL4 missense alleles provides insights into its ability to process distinct classes of dsRNA substrates , 2018, The Plant journal : for cell and molecular biology.

[28]  S. He,et al.  Pseudomonas syringae: what it takes to be a pathogen , 2018, Nature Reviews Microbiology.

[29]  Mark N. Puttick,et al.  The timescale of early land plant evolution , 2018, Proceedings of the National Academy of Sciences.

[30]  Kengo Kinoshita,et al.  ATTED-II in 2018: A Plant Coexpression Database Based on Investigation of the Statistical Property of the Mutual Rank Index , 2018, Plant & cell physiology.

[31]  Y. Kluger,et al.  Nlrp9b inflammasome restricts rotavirus infection in intestinal epithelial cells , 2017, Nature.

[32]  H. Vaucheret,et al.  DCL2‐ and RDR6‐dependent transitive silencing of SMXL4 and SMXL5 in Arabidopsis dcl4 mutants causes defective phloem transport and carbohydrate over‐accumulation , 2017, The Plant journal : for cell and molecular biology.

[33]  J. Agustí,et al.  Strigolactone- and Karrikin-Independent SMXL Proteins Are Central Regulators of Phloem Formation , 2017, Current Biology.

[34]  J. Mundy,et al.  Matching NLR Immune Receptors to Autoimmunity in camta3 Mutants Using Antimorphic NLR Alleles. , 2017, Cell host & microbe.

[35]  Qing Kong,et al.  The NLR protein SUMM2 senses the disruption of an immune signaling MAP kinase cascade via CRCK3 , 2017, EMBO reports.

[36]  O. Voinnet,et al.  DNA Methylation Influences the Expression of DICER-LIKE4 Isoforms, Which Encode Proteins of Alternative Localization and Function , 2016, Plant Cell.

[37]  T. Boller,et al.  Double-stranded RNAs induce a pattern-triggered immune signaling pathway in plants. , 2016, The New phytologist.

[38]  B. Kobe,et al.  Multiple Domain Associations within the Arabidopsis Immune Receptor RPP1 Regulate the Activation of Programmed Cell Death , 2016, PLoS pathogens.

[39]  Sudhir Kumar,et al.  MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets. , 2016, Molecular biology and evolution.

[40]  Hongwei Guo,et al.  Suppression of endogenous gene silencing by bidirectional cytoplasmic RNA decay in Arabidopsis , 2015, Science.

[41]  H. Vaucheret,et al.  Respective contributions of Arabidopsis DCL2 and DCL4 to RNA silencing. , 2015, The Plant journal : for cell and molecular biology.

[42]  Xuemei Chen,et al.  The Exosome and Trans-Acting Small Interfering RNAs Regulate Cuticular Wax Biosynthesis during Arabidopsis Inflorescence Stem Development1[OPEN] , 2014, Plant Physiology.

[43]  G. Rätsch,et al.  Species-wide Genetic Incompatibility Analysis Identifies Immune Genes as Hot Spots of Deleterious Epistasis , 2014, Cell.

[44]  W. Huber,et al.  Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2 , 2014, Genome Biology.

[45]  O. Dong,et al.  HSP90s are required for NLR immune receptor accumulation in Arabidopsis. , 2014, The Plant journal : for cell and molecular biology.

[46]  Zhijian J. Chen,et al.  Innate immune sensing and signaling of cytosolic nucleic acids. , 2014, Annual review of immunology.

[47]  O. Voinnet,et al.  The Arabidopsis miR472-RDR6 Silencing Pathway Modulates PAMP- and Effector-Triggered Immunity through the Post-transcriptional Control of Disease Resistance Genes , 2014, PLoS pathogens.

[48]  Wei Shi,et al.  featureCounts: an efficient general purpose program for assigning sequence reads to genomic features , 2013, Bioinform..

[49]  E. Miska,et al.  A deletion polymorphism in the Caenorhabditis elegans RIG-I homolog disables viral RNA dicing and antiviral immunity , 2013, eLife.

[50]  Thomas R. Gingeras,et al.  STAR: ultrafast universal RNA-seq aligner , 2013, Bioinform..

[51]  M. Axtell,et al.  DICER-LIKE3 activity in Physcomitrella patens DICER-LIKE4 mutants causes severe developmental dysfunction and sterility. , 2012, Molecular plant.

[52]  D. Higgins,et al.  Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega , 2011, Molecular systems biology.

[53]  Karsten M. Borgwardt,et al.  Whole-genome sequencing of multiple Arabidopsis thaliana populations , 2011, Nature Genetics.

[54]  Peter R. Crane,et al.  Early Flowers and Angiosperm Evolution , 2011 .

[55]  J. Dangl,et al.  Expanded functions for a family of plant intracellular immune receptors beyond specific recognition of pathogen effectors , 2011, Proceedings of the National Academy of Sciences.

[56]  Marcel Martin.  Cutadapt removes adapter sequences from high-throughput sequencing reads , 2011 .

[57]  H. Vaucheret,et al.  The 21-Nucleotide, but Not 22-Nucleotide, Viral Secondary Small Interfering RNAs Direct Potent Antiviral Defense by Two Cooperative Argonautes in Arabidopsis thaliana[W][OA] , 2011, Plant Cell.

[58]  William Stafford Noble,et al.  FIMO: scanning for occurrences of a given motif , 2011, Bioinform..

[59]  H. Nielsen,et al.  Autoimmunity in Arabidopsis acd11 Is Mediated by Epigenetic Regulation of an Immune Receptor , 2010, PLoS pathogens.

[60]  C. Glass,et al.  Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. , 2010, Molecular cell.

[61]  C. Sullivan,et al.  Arabidopsis RNA-Dependent RNA Polymerases and Dicer-Like Proteins in Antiviral Defense and Small Interfering RNA Biogenesis during Turnip Mosaic Virus Infection[W][OA] , 2010, Plant Cell.

[62]  S. Ding,et al.  An RIG-I-Like RNA Helicase Mediates Antiviral RNAi Downstream of Viral siRNA Biogenesis in Caenorhabditis elegans , 2009, PLoS pathogens.

[63]  A. Budd,et al.  The DExD/H-box helicase Dicer-2 mediates the induction of antiviral activity in drosophila , 2008, Nature Immunology.

[64]  F. Qu,et al.  Arabidopsis DRB4, AGO1, AGO7, and RDR6 participate in a DCL4-initiated antiviral RNA silencing pathway negatively regulated by DCL1 , 2008, Proceedings of the National Academy of Sciences.

[65]  Xuemei Chen,et al.  DICER-LIKE2 Plays a Primary Role in Transitive Silencing of Transgenes in Arabidopsis , 2008, PloS one.

[66]  Olivier Voinnet,et al.  Antiviral Immunity Directed by Small RNAs , 2007, Cell.

[67]  Gurman Singh Pall,et al.  Carbodiimide-mediated cross-linking of RNA to nylon membranes improves the detection of siRNA, miRNA and piRNA by northern blot , 2007, Nucleic acids research.

[68]  Jonathan D. G. Jones,et al.  The plant immune system , 2006, Nature.

[69]  R. Andino,et al.  The RNA silencing endonuclease Argonaute 2 mediates specific antiviral immunity in Drosophila melanogaster. , 2006, Genes & development.

[70]  Nicolas Bouché,et al.  An antagonistic function for Arabidopsis DCL2 in development and a new function for DCL4 in generating viral siRNAs , 2006, The EMBO journal.

[71]  Jinsong Bao,et al.  Hierarchical Action and Inhibition of Plant Dicer-Like Proteins in Antiviral Defense , 2006, Science.

[72]  R. Carthew,et al.  Methods and Materials , 1956, Eco-Art Therapy in Practice.

[73]  A. Schneemann,et al.  Essential function in vivo for Dicer-2 in host defense against RNA viruses in drosophila , 2006, Nature Immunology.

[74]  Thomas Lengauer,et al.  Mutations in the NB-ARC Domain of I-2 That Impair ATP Hydrolysis Cause Autoactivation1[OA] , 2006, Plant Physiology.

[75]  A. Bent.  Faculty Opinions recommendation of Elicitor-mediated oligomerization of the tobacco N disease resistance protein. , 2006 .

[76]  Edwards Allen,et al.  DICER-LIKE 4 functions in trans-acting small interfering RNA biogenesis and vegetative phase change in Arabidopsis thaliana. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[77]  David P. Bartel,et al.  Partially Redundant Functions of Arabidopsis DICER-like Enzymes and a Role for DCL4 in Producing trans-Acting siRNAs , 2005, Current Biology.

[78]  J. Dangl,et al.  Antagonistic Control of Disease Resistance Protein Stability in the Plant Immune System , 2005, Science.

[79]  Imre E Somssich,et al.  WRKY transcription factors: from DNA binding towards biological function. , 2004, Current opinion in plant biology.

[80]  J. Beynon,et al.  The arabidopsis TIR-NB-LRR gene RAC1 confers resistance to Albugo candida (white rust) and is dependent on EDS1 but not PAD4. , 2004, Molecular plant-microbe interactions : MPMI.

[81]  Adam M. Gustafson,et al.  Genetic and Functional Diversification of Small RNA Pathways in Plants , 2004, PLoS biology.

[82]  N. Ashton,et al.  The isolation and preliminary characterisation of auxotrophic and analogue resistant mutants of the moss, Physcomitrella patens , 1977, Molecular and General Genetics MGG.

[83]  J. Dangl,et al.  Cytosolic HSP90 associates with and modulates the Arabidopsis RPM1 disease resistance protein , 2003, The EMBO journal.

[84]  K. Shirasu,et al.  HSP90 interacts with RAR1 and SGT1 and is essential for RPS2-mediated disease resistance in Arabidopsis , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[85]  Jack E. Dixon,et al.  Cleavage of Arabidopsis PBS1 by a Bacterial Type III Effector , 2003, Science.

[86]  Michael J. Axtell,et al.  Initiation of RPS2-Specified Disease Resistance in Arabidopsis Is Coupled to the AvrRpt2-Directed Elimination of RIN4 , 2003, Cell.

[87]  J. Vossen,et al.  The Tomato R Gene Products I-2 and Mi-1 Are Functional ATP Binding Proteins with ATPase Activity , 2002, The Plant Cell Online.

[88]  Jonathan D. G. Jones,et al.  Ubiquitin ligase-associated protein SGT1 is required for host and nonhost disease resistance in plants , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[89]  T. Eulgem,et al.  Arabidopsis SGT1b Is Required for Defense Signaling Conferred by Several Downy Mildew Resistance Genes Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.001123. , 2002, The Plant Cell Online.

[90]  Jonathan D. G. Jones,et al.  A Tomato Cysteine Protease Required for Cf-2-Dependent Disease Resistance and Suppression of Autonecrosis , 2002, Science.

[91]  Ken Shirasu,et al.  The RAR1 Interactor SGT1, an Essential Component of R Gene-Triggered Disease Resistance , 2002, Science.

[92]  Jonathan D. G. Jones,et al.  Regulatory Role of SGT1 in Early R Gene-Mediated Plant Defenses , 2002, Science.

[93]  Jonathan D. G. Jones,et al.  Plant pathogens and integrated defence responses to infection , 2001, Nature.

[94]  J. Beynon,et al.  White rust (Albugo candida) resistance loci on three Arabidopsis chromosomes are closely linked to downy mildew (Peronospora parasitica) resistance loci. , 2001, Molecular plant pathology.

[95]  Erik Andreasson,et al.  Arabidopsis MAP Kinase 4 Negatively Regulates Systemic Acquired Resistance , 2000, Cell.

[96]  T. Eulgem,et al.  The transcriptome of Arabidopsis thaliana during systemic acquired resistance , 2000, Nature Genetics.

[97]  G. Neuhaus,et al.  Nuclear export of proteins in plants: AtXPO1 is the export receptor for leucine-rich nuclear export signals in Arabidopsis thaliana. , 1999, The Plant journal : for cell and molecular biology.

[98]  S. Jacobsen,et al.  Disruption of an RNA helicase/RNAse III gene in Arabidopsis causes unregulated cell division in floral meristems. , 1999, Development.

[99]  D. Klessig,et al.  The Arabidopsis ssi1 Mutation Restores Pathogenesis-Related Gene Expression in npr1 Plants and Renders Defensin Gene Expression Salicylic Acid Dependent , 1999, Plant Cell.

[100]  E. A. van der Biezen,et al.  Plant disease-resistance proteins and the gene-for-gene concept. , 1998, Trends in biochemical sciences.

[101]  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.

[102]  J Chory,et al.  dCAPS, a simple technique for the genetic analysis of single nucleotide polymorphisms: experimental applications in Arabidopsis thaliana genetics. , 1998, The Plant journal : for cell and molecular biology.

[103]  Leslie Friedrich,et al.  Requirement of Salicylic Acid for the Induction of Systemic Acquired Resistance , 1993, Science.

[104]  N. Saitou,et al.  The neighbor-joining method: a new method for reconstructing phylogenetic trees. , 1987, Molecular biology and evolution.

[105]  J. Felsenstein.  CONFIDENCE LIMITS ON PHYLOGENIES: AN APPROACH USING THE BOOTSTRAP , 1985, Evolution; international journal of organic evolution.

[106]  L. Pauling,et al.  Molecules as documents of evolutionary history. , 1965, Journal of theoretical biology.

[107]  Thomas D. Schmittgen,et al.  Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2 2 DD C T Method , 2022 .