Virus-induced gene silencing offers a functional genomics platform for studying plant cell wall formation.

Virus-induced gene silencing (VIGS) is a powerful genetic tool for rapid assessment of plant gene functions in the post-genomic era. Here, we successfully implemented a Tobacco Rattle Virus (TRV)-based VIGS system to study functions of genes involved in either primary or secondary cell wall formation in Nicotiana benthamiana plants. A 3-week post-VIGS time frame is sufficient to observe phenotypic alterations in the anatomical structure of stems and chemical composition of the primary and secondary cell walls. We used cell wall glycan-directed monoclonal antibodies to demonstrate that alteration of cell wall polymer synthesis during the secondary growth phase of VIGS plants has profound effects on the extractability of components from woody stem cell walls. Therefore, TRV-based VIGS together with cell wall component profiling methods provide a high-throughput gene discovery platform for studying plant cell wall formation from a bioenergy perspective.

[1]  William S York,et al.  A Comprehensive Toolkit of Plant Cell Wall Glycan-Directed Monoclonal Antibodies1[W][OA] , 2010, Plant Physiology.

[2]  G. Volckaert,et al.  Plant cell walls: Protecting the barrier from degradation by microbial enzymes. , 2009, Seminars in cell & developmental biology.

[3]  G. S. Randhawa,et al.  Plant cell wall matrix polysaccharide biosynthesis. , 2009, Molecular plant.

[4]  C. Somerville,et al.  Cell Wall Polysaccharide Synthesis , 2009 .

[5]  William S York,et al.  Biochemical control of xylan biosynthesis - which end is up? , 2008, Current opinion in plant biology.

[6]  W. Willats,et al.  Pectic homogalacturonan masks abundant sets of xyloglucan epitopes in plant cell walls , 2008, BMC Plant Biology.

[7]  Markus Pauly,et al.  Cell-wall carbohydrates and their modification as a resource for biofuels. , 2008, The Plant journal : for cell and molecular biology.

[8]  R. Goodacre,et al.  Comparison of five xylan synthesis mutants reveals new insight into the mechanisms of xylan synthesis. , 2007, The Plant journal : for cell and molecular biology.

[9]  S. Dinesh-Kumar,et al.  A Ligation-Independent Cloning Tobacco Rattle Virus Vector for High-Throughput Virus-Induced Gene Silencing Identifies Roles for NbMADS4-1 and -2 in Floral Development1[W][OA] , 2007, Plant Physiology.

[10]  O. Vandeputte,et al.  From primary to secondary growth: origin and development of the vascular system. , 2007, Journal of experimental botany.

[11]  Antony Bacic,et al.  High-throughput mapping of cell-wall polymers within and between plants using novel microarrays. , 2007, The Plant journal : for cell and molecular biology.

[12]  R. Zhong,et al.  Arabidopsis irregular xylem8 and irregular xylem9: Implications for the Complexity of Glucuronoxylan Biosynthesis[W] , 2007, The Plant Cell Online.

[13]  Chris Somerville,et al.  Cellulose synthesis in higher plants. , 2006, Annual review of cell and developmental biology.

[14]  D. Bolam,et al.  Understanding the Biological Rationale for the Diversity of Cellulose-directed Carbohydrate-binding Modules in Prokaryotic Enzymes* , 2006, Journal of Biological Chemistry.

[15]  S. Jayanty,et al.  Loss of function of COBRA, a determinant of oriented cell expansion, invokes cellular defence responses in Arabidopsis thaliana. , 2006, Journal of experimental botany.

[16]  R. Meeley,et al.  Brittle stalk 2 encodes a putative glycosylphosphatidylinositol-anchored protein that affects mechanical strength of maize tissues by altering the composition and structure of secondary cell walls , 2006, Planta.

[17]  Antony Bacic,et al.  Plant cell wall biosynthesis: genetic, biochemical and functional genomics approaches to the identification of key genes. , 2006, Plant biotechnology journal.

[18]  D. Cosgrove Growth of the plant cell wall , 2005, Nature Reviews Molecular Cell Biology.

[19]  T. Baskin Anisotropic expansion of the plant cell wall. , 2005, Annual review of cell and developmental biology.

[20]  Maureen C. McCann,et al.  Genomics of plant cell wall biogenesis , 2005, Planta.

[21]  Royston Goodacre,et al.  Identification of Novel Genes in Arabidopsis Involved in Secondary Cell Wall Formation Using Expression Profiling and Reverse Genetics , 2005, The Plant Cell Online.

[22]  Staffan Persson,et al.  Identification of genes required for cellulose synthesis by regression analysis of public microarray data sets. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[23]  P. Benfey,et al.  COBRA, an Arabidopsis Extracellular Glycosyl-Phosphatidyl Inositol-Anchored Protein, Specifically Controls Highly Anisotropic Expansion through Its Involvement in Cellulose Microfibril Orientationw⃞ , 2005, The Plant Cell Online.

[24]  S. Nishimura,et al.  Carbohydrate analysis by a phenol-sulfuric acid method in microplate format. , 2005, Analytical biochemistry.

[25]  J. Knox,et al.  Monoclonal Antibodies to Plant Cell Wall Xylans and Arabinoxylans , 2005, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[26]  B. Sundberg,et al.  Carbohydrate-Active Enzymes Involved in the Secondary Cell Wall Biogenesis in Hybrid Aspen1 , 2005, Plant Physiology.

[27]  U. Kalluri,et al.  Genomics of cellulose biosynthesis in poplars. , 2004, The New phytologist.

[28]  B. Sundberg,et al.  A Populus EST resource for plant functional genomics. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[29]  G. Martin,et al.  Applications and advantages of virus-induced gene silencing for gene function studies in plants. , 2004, The Plant journal : for cell and molecular biology.

[30]  D. Robertson VIGS vectors for gene silencing: many targets, many tools. , 2004, Annual review of plant biology.

[31]  Q. Qian,et al.  BRITTLE CULM1, Which Encodes a COBRA-Like Protein, Affects the Mechanical Properties of Rice Plants Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.011775. , 2003, The Plant Cell Online.

[32]  David C Baulcombe,et al.  Virus-induced gene silencing in plants. , 2003, Methods.

[33]  N. Lewis,et al.  Trends in lignin modification: a comprehensive analysis of the effects of genetic manipulations/mutations on lignification and vascular integrity. , 2002, Phytochemistry.

[34]  S. Dinesh-Kumar,et al.  Tobacco Rar1, EDS1 and NPR1/NIM1 like genes are required for N-mediated resistance to tobacco mosaic virus. , 2002, The Plant journal : for cell and molecular biology.

[35]  Jacqueline Grima-Pettenati,et al.  Down-regulation of Cinnamoyl-CoA reductase induces significant changes of lignin profiles in transgenic tobacco plants , 2002 .

[36]  J. Grima-Pettenati,et al.  Strong decrease in lignin content without significant alteration of plant development is induced by simultaneous down-regulation of cinnamoyl CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) in tobacco plants. , 2001, The Plant journal : for cell and molecular biology.

[37]  Björn Sundberg,et al.  Unravelling cell wall formation in the woody dicot stem , 2001, Plant Molecular Biology.

[38]  P. Benfey,et al.  COBRA encodes a putative GPI-anchored protein, which is polarly localized and necessary for oriented cell expansion in Arabidopsis. , 2001, Genes & development.

[39]  S. Turner,et al.  Multiple Cellulose Synthase Catalytic Subunits Are Required for Cellulose Synthesis in Arabidopsis , 2000, Plant Cell.

[40]  Guislaine Refregier,et al.  PROCUSTE1 Encodes a Cellulose Synthase Required for Normal Cell Elongation Specifically in Roots and Dark-Grown Hypocotyls of Arabidopsis , 2000, Plant Cell.

[41]  Rachel A. Burton,et al.  Virus-Induced Silencing of a Plant Cellulose Synthase Gene , 2000, Plant Cell.

[42]  R. Zhong,et al.  Dual Methylation Pathways in Lignin Biosynthesis , 1998, Plant Cell.

[43]  M. Van Montagu,et al.  Gene discovery in the wood-forming tissues of poplar: analysis of 5, 692 expressed sequence tags. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[44]  Y. Katayama,et al.  Immunological characterization of transgenic tobacco plants with a chimeric gene for 4-coumarate:CoA ligase that have altered lignin in their xylem tissue , 1997 .

[45]  Bernard Fritig,et al.  Altered lignin composition in transgenic tobacco expressing O-methyltransferase sequences in sense and antisense orientation , 1995 .

[46]  R. Dixon,et al.  Reduced lignin in transgenic plants containing a caffeic acidO-methyltransferase antisense gene , 1994, Transgenic Research.

[47]  B. Stone,et al.  The application of Sirofluor, a chemically defined fluorochrome from aniline blue for the histochemical detection of callose , 1984, Protoplasma.

[48]  B. Stone,et al.  Studies on the specificity of interaction of cereal cell wall components with Congo Red and Calcofluor. Specific detection and histochemistry of (1→3),(1→4),-β-D-glucan , 1983 .

[49]  P. Ahlgren,et al.  Removal of Wood Components During Chlorite Delignification of Black Spruce , 1971 .

[50]  S. Dinesh-Kumar,et al.  Virus-induced gene silencing as a tool to identify host genes affecting viral pathogenicity. , 2008, Methods in molecular biology.