title: Functional dissection of a strong and specific microbe-associated molecular pattern-responsive synthetic promoter.
abstract: Synthetic promoters are important for temporal and spatial gene expression in transgenic plants. To identify novel microbe-associated molecular pattern (MAMP)-responsive cis-regulatory sequences for synthetic promoter design, a combination of bioinformatics and experimental approaches was employed. One cis-sequence was identified which confers strong MAMP-responsive reporter gene activity with low background activity. The 35-bp-long cis-sequence was identified in the promoter of the Arabidopsis thaliana DJ1E gene, a homologue of the human oncogene DJ1. In this study, this cis-sequence is shown to be a tripartite cis-regulatory module (CRM). A synthetic promoter with four copies of the CRM linked to a minimal promoter increases MAMP-responsive reporter gene expression compared to the wild-type DJ1E promoter. The CRM consists of two WT-boxes (GGACTTTT and GGACTTTG) and a variant of the GCC-box (GCCACC), all required for MAMP and salicylic acid (SA) responsivity. Yeast one-hybrid screenings using a transcription factor (TF)-only prey library identified two AP2/ERFs, ORA59 and ERF10, interacting antagonistically with the CRM. ORA59 activates reporter gene activity and requires the consensus core sequence GCCNCC for gene expression activation. ERF10 down-regulates MAMP-responsive gene expression. No TFs interacting with the WT-boxes GGACTTTT and GGACTTTG were selected in yeast one-hybrid screenings with the TF-only prey library. In transgenic Arabidopsis, the synthetic promoter confers strong and specific reporter gene activity in response to biotrophs and necrotrophs as well as SA.
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Mona
authors.initials: M
authors.lastname: Lehmeyer
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Konstantin
authors.initials: K
authors.lastname: Kanofsky
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Erik K R
authors.initials: EK
authors.lastname: Hanko
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Ahrendt
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Maren
authors.initials: M
authors.lastname: Wehrs
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Machens
authors.affiliation: Institut für Genetik, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Hehl
mesh_terms: D017360:Arabidopsis / Q000187:drug effects / Q000235:genetics / Q000382:microbiology; D018744:DNA, Plant / Q000235:genetics* / Q000378:metabolism; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D053263:Gene Regulatory Networks / Q000187:drug effects; D017343:Genes, Plant; D017930:Genes, Reporter; D000069452:Pathogen-Associated Molecular Pattern Molecules / Q000378:metabolism*; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D020156:Salicylic Acid / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D018744:DNA, Plant; D000069452:Pathogen-Associated Molecular Pattern Molecules; D014157:Transcription Factors; D020156:Salicylic Acid
keywords: plant synthetic biology; plant-pathogen interaction; transgenic plants; transient reporter gene assays
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