title: Dirigent Protein-Mediated Lignan and Cyanogenic Glucoside Formation in Flax Seed: Integrated Omics and MALDI Mass Spectrometry Imaging.
abstract: An integrated omics approach using genomics, transcriptomics, metabolomics (MALDI mass spectrometry imaging, MSI), and bioinformatics was employed to study spatiotemporal formation and deposition of health-protecting polymeric lignans and plant defense cyanogenic glucosides. Intact flax (Linum usitatissimum) capsules and seed tissues at different development stages were analyzed. Transcriptome analyses indicated distinct expression patterns of dirigent protein (DP) gene family members encoding (-)- and (+)-pinoresinol-forming DPs and their associated downstream metabolic processes, respectively, with the former expressed at early seed coat development stages. Genes encoding (+)-pinoresinol-forming DPs were, in contrast, expressed at later development stages. Recombinant DP expression and DP assays also unequivocally established their distinct stereoselective biochemical functions. Using MALDI MSI and ion mobility separation analyses, the pinoresinol downstream derivatives, secoisolariciresinol diglucoside (SDG) and SDG hydroxymethylglutaryl ester, were localized and detectable only in early seed coat development stages. SDG derivatives were then converted into higher molecular weight phenolics during seed coat maturation. By contrast, the plant defense cyanogenic glucosides, the monoglucosides linamarin/lotaustralin, were detected throughout the flax capsule, whereas diglucosides linustatin/neolinustatin only accumulated in endosperm and embryo tissues. A putative biosynthetic pathway to the cyanogens is proposed on the basis of transcriptome coexpression data. Localization of all metabolites was at ca. 20 μm resolution, with the web based tool OpenMSI enabling not only resolution enhancement but also an interactive system for real-time searching for any ion in the tissue under analysis.
authors.affiliation: †Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, United States.
authors.forename: Doralyn S
authors.initials: DS
authors.lastname: Dalisay
authors.affiliation: †Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, United States.
authors.forename: Kye Won
authors.initials: KW
authors.lastname: Kim
authors.affiliation: †Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, United States.
authors.forename: Choonseok
authors.initials: C
authors.lastname: Lee
authors.affiliation: †Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, United States.
authors.forename: Hong
authors.initials: H
authors.lastname: Yang
authors.affiliation: ‡Computational Research Division, Lawrence Berkeley National Laboratory, One Cyclotron Road, Berkeley, California 94720, United States.
authors.forename: Oliver
authors.initials: O
authors.lastname: Rübel
authors.affiliation: §Life Sciences Division, Lawrence Berkeley National Laboratory, One Cyclotron Road, Berkeley, California 94720, United States.
authors.forename: Benjamin P
authors.initials: BP
authors.lastname: Bowen
authors.affiliation: †Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, United States.
authors.forename: Laurence B
authors.initials: LB
authors.lastname: Davin
authors.affiliation: †Institute of Biological Chemistry, Washington State University, Pullman, Washington 99164-6340, United States.
authors.forename: Norman G
authors.initials: NG
authors.lastname: Lewis
mesh_terms: D002072:Butylene Glycols / Q000032:analysis; D019597:Flax / Q000737:chemistry* / Q000235:genetics; D005663:Furans / Q000032:analysis / Q000737:chemistry*; D005960:Glucosides / Q000032:analysis; D006027:Glycosides / Q000032:analysis / Q000737:chemistry*; D017705:Lignans / Q000032:analysis / Q000737:chemistry*; D015394:Molecular Structure; D009570:Nitriles / Q000032:analysis; D012639:Seeds / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D002072:Butylene Glycols; D005663:Furans; D005960:Glucosides; D006027:Glycosides; D017705:Lignans; D009570:Nitriles; C060283:secoisolariciresinol; C001556:lotaustralin; C090142:secoisolariciresinol diglucoside; C103298:pinoresinol