3
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: found
      Is Open Access

      CRISPR/Cas9-Induced fad2 and rod1 Mutations Stacked With fae1 Confer High Oleic Acid Seed Oil in Pennycress ( Thlaspi arvense L.)

      research-article

      Read this article at

      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Pennycress ( Thlaspi arvense L.) is being domesticated as an oilseed cash cover crop to be grown in the off-season throughout temperate regions of the world. With its diploid genome and ease of directed mutagenesis using molecular approaches, pennycress seed oil composition can be rapidly tailored for a plethora of food, feed, oleochemical and fuel uses. Here, we utilized Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 technology to produce knockout mutations in the FATTY ACID DESATURASE2 ( FAD2) and REDUCED OLEATE DESATURATION1 ( ROD1) genes to increase oleic acid content. High oleic acid (18:1) oil is valued for its oxidative stability that is superior to the polyunsaturated fatty acids (PUFAs) linoleic (18:2) and linolenic (18:3), and better cold flow properties than the very long chain fatty acid (VLCFA) erucic (22:1). When combined with a FATTY ACID ELONGATION1 ( fae1) knockout mutation, fad2 fae1 and rod1 fae1 double mutants produced ∼90% and ∼60% oleic acid in seed oil, respectively, with PUFAs in fad2 fae1 as well as fad2 single mutants reduced to less than 5%. MALDI-MS spatial imaging analyses of phosphatidylcholine (PC) and triacylglycerol (TAG) molecular species in wild-type pennycress embryo sections from mature seeds revealed that erucic acid is highly enriched in cotyledons which serve as storage organs, suggestive of a role in providing energy for the germinating seedling. In contrast, PUFA-containing TAGs are enriched in the embryonic axis, which may be utilized for cellular membrane expansion during seed germination and seedling emergence. Under standard growth chamber conditions, rod1 fae1 plants grew like wild type whereas fad2 single and fad2 fae1 double mutant plants exhibited delayed growth and overall reduced heights and seed yields, suggesting that reducing PUFAs below a threshold in pennycress had negative physiological effects. Taken together, our results suggest that combinatorial knockout of ROD1 and FAE1 may be a viable route to commercially increase oleic acid content in pennycress seed oil whereas mutations in FAD2 will likely require at least partial function to avoid fitness trade-offs.

          Related collections

          Most cited references103

          • Record: found
          • Abstract: found
          • Article: not found

          Both CRISPR/Cas-based nucleases and nickases can be used efficiently for genome engineering in Arabidopsis thaliana.

          Engineered nucleases can be used to induce site-specific double-strand breaks (DSBs) in plant genomes. Thus, homologous recombination (HR) can be enhanced and targeted mutagenesis can be achieved by error-prone non-homologous end-joining (NHEJ). Recently, the bacterial CRISPR/Cas9 system was used for DSB induction in plants to promote HR and NHEJ. Cas9 can also be engineered to work as a nickase inducing single-strand breaks (SSBs). Here we show that only the nuclease but not the nickase is an efficient tool for NHEJ-mediated mutagenesis in plants. We demonstrate the stable inheritance of nuclease-induced targeted mutagenesis events in the ADH1 and TT4 genes of Arabidopsis thaliana at frequencies from 2.5 up to 70.0%. Deep sequencing analysis revealed NHEJ-mediated DSB repair in about a third of all reads in T1 plants. In contrast, applying the nickase resulted in the reduction of mutation frequency by at least 740-fold. Nevertheless, the nickase is able to induce HR at similar efficiencies as the nuclease or the homing endonuclease I-SceI. Two different types of somatic HR mechanisms, recombination between tandemly arranged direct repeats as well as gene conversion using the information on an inverted repeat could be enhanced by the nickase to a similar extent as by DSB-inducing enzymes. Thus, the Cas9 nickase has the potential to become an important tool for genome engineering in plants. It should not only be applicable for HR-mediated gene targeting systems but also by the combined action of two nickases as DSB-inducing agents excluding off-target effects in homologous genomic regions. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.
            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            Jasmonate action in plant growth and development.

            Phytohormones, including jasmonates (JAs), gibberellin, ethylene, abscisic acid, and auxin, integrate endogenous developmental cues with environmental signals to regulate plant growth, development, and defense. JAs are well- recognized lipid-derived stress hormones that regulate plant adaptations to biotic stresses, including herbivore attack and pathogen infection, as well as abiotic stresses, including wounding, ozone, and ultraviolet radiation. An increasing number of studies have shown that JAs also have functions in a remarkable number of plant developmental events, including primary root growth, reproductive development, and leaf senescence. Since the 1980s, details of the JA biosynthesis pathway, signaling pathway, and crosstalk during plant growth and development have been elucidated. Here, we summarize recent advances and give an updated overview of JA action and crosstalk in plant growth and development.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: not found

              Compartmentation of triacylglycerol accumulation in plants.

              Triacylglycerols from plants, familiar to most people as vegetable oils, supply 25% of dietary calories to the developed world and are increasingly a source for renewable biomaterials and fuels. Demand for vegetable oils will double by 2030, which can be met only by increased oil production. Triacylglycerol synthesis is accomplished through the coordinate action of multiple pathways in multiple subcellular compartments. Recent information has revealed an underappreciated complexity in pathways for synthesis and accumulation of this important energy-rich class of molecules.
                Bookmark

                Author and article information

                Contributors
                Journal
                Front Plant Sci
                Front Plant Sci
                Front. Plant Sci.
                Frontiers in Plant Science
                Frontiers Media S.A.
                1664-462X
                22 April 2021
                2021
                : 12
                : 652319
                Affiliations
                [1] 1School of Biological Sciences, Illinois State University , Normal, IL, United States
                [2] 2BioDiscovery Institute and Department of Biological Sciences, University of North Texas , Denton, TX, United States
                [3] 3Department of Biochemistry and Center for Plant Science Innovation, University of Nebraska-Lincoln , Lincoln, NE, United States
                Author notes

                Edited by: Juliette Jouhet, UMR 5168 Laboratoire de Physiologie Cellulaire Vegetale (LPCV), France

                Reviewed by: José Manuel Martínez-Rivas, Instituto de la Grasa (CSIC), Spain; Clay Carter, University of Minnesota Twin Cities, United States

                *Correspondence: John C. Sedbrook, jcsedbr@ 123456ilstu.edu

                This article was submitted to Plant Physiology, a section of the journal Frontiers in Plant Science

                Article
                10.3389/fpls.2021.652319
                8100250
                33968108
                c372ee1d-3d2b-4371-9d77-51d28529b881
                Copyright © 2021 Jarvis, Romsdahl, McGinn, Nazarenus, Cahoon, Chapman and Sedbrook.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 12 January 2021
                : 23 March 2021
                Page count
                Figures: 8, Tables: 0, Equations: 0, References: 104, Pages: 18, Words: 0
                Funding
                Funded by: U.S. Department of Agriculture 10.13039/100000199
                Award ID: 2018-67009-27374
                Award ID: 2019-69012-29851
                Award ID: 2018-38821-28111
                Categories
                Plant Science
                Original Research

                Plant science & Botany
                crispr,maldi-msi,oilseed,oleic acid,pennycress,polyunsaturated fatty acid,thlaspi arvense,triacylglycerol

                Comments

                Comment on this article