title: An engineered promoter driving expression of a microbial avirulence gene confers recognition of TAL effectors and reduces growth of diverse Xanthomonas strains in citrus.
abstract: Xanthomonas citri ssp. citri (X. citri), causal agent of citrus canker, uses transcription activator-like effectors (TALEs) as major pathogenicity factors. TALEs, which are delivered into plant cells through the type III secretion system (T3SS), interact with effector binding elements (EBEs) in host genomes to activate the expression of downstream susceptibility genes to promote disease. Predictably, TALEs bind EBEs in host promoters via known combinations of TALE amino acids to DNA bases, known as the TALE code. We introduced 14 EBEs, matching distinct X. citri TALEs, into the promoter of the pepper Bs3 gene (ProBs31EBE), and fused this engineered promoter with multiple EBEs (ProBs314EBE) to either the β-glucuronidase (GUS) reporter gene or the coding sequence (cds) of the pepper gene, Bs3. TALE-induced expression of the Bs3 cds in citrus leaves resulted in no visible hypersensitive response (HR). Therefore, we utilized a different approach in which ProBs31EBEand ProBs314EBEwere fused to the Xanthomonas gene, avrGf1, which encodes a bacterial effector that elicits an HR in grapefruit and sweet orange. We demonstrated, in transient assays, that activation of ProBs314EBEby X. citri TALEs is T3SS dependent, and that the expression of AvrGf1 triggers HR and correlates with reduced bacterial growth. We further demonstrated that all tested virulent X. citri strains from diverse geographical locations activate ProBs314EBE. TALEs are essential for the virulence of X. citri strains and, because the engineered promoter traps are activated by multiple TALEs, this concept has the potential to confer broad-spectrum, durable resistance to citrus canker in stably transformed plants.
authors.affiliation: Plant Pathology Department, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Deepak
authors.initials: D
authors.lastname: Shantharaj
authors.affiliation: Genetics, Department of Biology, Ludwig-Maximilians-University Munich, Martinsried, D-82152, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Römer
authors.affiliation: Plant Pathology Department, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Jose F L
authors.initials: JFL
authors.lastname: Figueiredo
authors.affiliation: Plant Pathology Department, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Gerald V
authors.initials: GV
authors.lastname: Minsavage
authors.affiliation: Zentrum für Molekularbiologie der Pflanzen (ZMBP), Eberhard-Karls-Universität Tübingen, Auf der Morgenstelle 32, Tübingen, D-72076, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Krönauer
authors.affiliation: Plant Pathology Department, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Stall
authors.affiliation: Department of Horticultural Sciences, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Gloria A
authors.initials: GA
authors.lastname: Moore
authors.affiliation: Department of Horticultural Sciences, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Latanya C
authors.initials: LC
authors.lastname: Fisher
authors.affiliation: Plant Pathology Department, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Hu
authors.affiliation: 2Blades Foundation, Suite 1901, 1630 Chicago Avenue, Evanston, IL, 60201, USA.
authors.forename: Diana M
authors.initials: DM
authors.lastname: Horvath
authors.affiliation: Genetics, Department of Biology, Ludwig-Maximilians-University Munich, Martinsried, D-82152, Germany.|Zentrum für Molekularbiologie der Pflanzen (ZMBP), Eberhard-Karls-Universität Tübingen, Auf der Morgenstelle 32, Tübingen, D-72076, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Lahaye
authors.affiliation: Plant Pathology Department, University of Florida, Gainesville, FL 32611, USA.
authors.forename: Jeffrey B
authors.initials: JB
authors.lastname: Jones
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002212:Capsicum / Q000235:genetics; D016923:Cell Death; D002957:Citrus / Q000382:microbiology*; D003429:Cross Reactions; D018506:Gene Expression Regulation, Plant; D005798:Genes, Bacterial; D005818:Genetic Engineering; D009154:Mutation / Q000235:genetics; D018515:Plant Leaves / Q000382:microbiology; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D000071336:Transcription Activator-Like Effectors / Q000378:metabolism*; D014774:Virulence / Q000235:genetics; D014974:Xanthomonas / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity*
chemical_list: D001426:Bacterial Proteins; D000071336:Transcription Activator-Like Effectors
keywords: avrGf1; effector binding element (EBE); hypersensitive response (HR); pathogen-inducible promoter; transcription activator-like effector (TALE)