title: Proteomic Identification of Putative MicroRNA394 Target Genes in Arabidopsis thaliana Identifies Major Latex Protein Family Members Critical for Normal Development.
abstract: Expression of the F-Box protein Leaf Curling Responsiveness (LCR) is regulated by microRNA, miR394, and alterations to this interplay in Arabidopsis thaliana produce defects in leaf polarity and shoot apical meristem organization. Although the miR394-LCR node has been documented in Arabidopsis, the identification of proteins targeted by LCR F-box itself has proven problematic. Here, a proteomic analysis of shoot apices from plants with altered LCR levels identified a member of the Latex Protein (MLP) family gene as a potential LCR F-box target. Bioinformatic and molecular analyses also suggested that other MLP family members are likely to be targets for this post-translational regulation. Direct interaction between LCR F-Box and MLP423 was validated. Additional MLP members had reduction in protein accumulation, in varying degrees, mediated by LCR F-Box. Transgenic Arabidopsis lines, in which MLP28 expression was reduced through an artificial miRNA technology, displayed severe developmental defects, including changes in leaf patterning and morphology, shoot apex defects, and eventual premature death. These phenotypic characteristics resemble those of Arabidopsis plants modified to over-express LCR Taken together, the results demonstrate that MLPs are driven to degradation by LCR, and indicate that MLP gene family is target of miR394-LCR regulatory node, representing potential targets for directly post-translational regulation mediated by LCR F-Box. In addition, MLP28 family member is associated with the LCR regulation that is critical for normal Arabidopsis development.
authors.affiliation: From the ‡School of Biological Sciences, The University of Sydney, Camperdown NSW 2006, Australia; §Laboratório de Biologia Molecular de Plantas, Universidade Federal do Rio de Janeiro, Cidade Universitária, RJ, Brazil; cgli2101@uni.sydney.edu.au.
authors.forename: Celso G
authors.initials: CG
authors.lastname: Litholdo
authors.affiliation: ¶Charles Perkins Centre, School of Molecular Bioscience, The University of Sydney, Darlington NSW 2006, Australia;
authors.forename: Benjamin L
authors.initials: BL
authors.lastname: Parker
authors.affiliation: ‖School of Environmental and Life Sciences, The University of Newcastle, Callaghan NSW 2308, Australia;
authors.forename: Andrew L
authors.initials: AL
authors.lastname: Eamens
authors.affiliation: **Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense 5230, Denmark;
authors.forename: Martin R
authors.initials: MR
authors.lastname: Larsen
authors.affiliation: ¶Charles Perkins Centre, School of Molecular Bioscience, The University of Sydney, Darlington NSW 2006, Australia;
authors.forename: Stuart J
authors.initials: SJ
authors.lastname: Cordwell
authors.affiliation: From the ‡School of Biological Sciences, The University of Sydney, Camperdown NSW 2006, Australia; ‡‡Centre for Tropical Crops and Biocommodities, Queensland University of Technology, Brisbane QLD 4001, Australia.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Waterhouse
mesh_terms: D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000378:metabolism*; D002853:Chromatography, Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D044783:F-Box Proteins; D018506:Gene Expression Regulation, Plant; D035683:MicroRNAs / Q000378:metabolism*; D005810:Multigene Family; D018520:Plant Shoots / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry
chemical_list: D029681:Arabidopsis Proteins; D044783:F-Box Proteins; C579342:MIRN394 microRNA, Arabidopsis; C542103:MLP28 protein, Arabidopsis; D035683:MicroRNAs; D003577:Cytochrome P-450 Enzyme System; C434730:CYP86A8 protein, Arabidopsis