OsNAC5 overexpression enlarges root diameter in rice plants leading to enhanced drought tolerance and increased grain yield in the field.

Drought conditions are among the most serious challenges to crop production worldwide. Here, we report the results of field evaluations of transgenic rice plants overexpressing OsNAC5, under the control of either the root-specific (RCc3) or constitutive (GOS2) promoters. Field evaluations over three growing seasons revealed that the grain yield of the RCc3:OsNAC5 and GOS2:OsNAC5 plants were increased by 9%-23% and 9%-26% under normal conditions, respectively. Under drought conditions, however, RCc3:OsNAC5 plants showed a significantly higher grain yield of 22%-63%, whilst the GOS2:OsNAC5 plants showed a reduced or similar yield to the nontransgenic (NT) controls. Both the RCc3:OsNAC5 and GOS2:OsNAC5 plants were found to have larger roots due to an enlarged stele and aerenchyma at flowering stage. Cell numbers per cortex layer and stele of developing roots were higher in both transgenic plants than NT controls, contributing to the increase in root diameter. The root diameter was enlarged to a greater extent in the RCc3:OsNAC5, suggesting the importance of this phenotype for enhanced drought tolerance. Microarray experiments identified 25 up-regulated genes by more than three-fold (P < 0.01) in the roots of both transgenic lines. Also identified were 19 and 18 up-regulated genes that are specific to the RCc3:OsNAC5 and GOS2:OsNAC5 roots, respectively. Of the genes specifically up-regulated in the RCc3:OsNAC5 roots, GLP, PDX, MERI5 and O-methyltransferase were implicated in root growth and development. Our present findings demonstrate that the root-specific overexpression of OsNAC5 enlarges roots significantly and thereby enhances drought tolerance and grain yield under field conditions.

[1]  J. Bailey-Serres,et al.  Posttranscriptional Control of Photosynthetic mRNA Decay under Stress Conditions Requires 3′ and 5′ Untranslated Regions and Correlates with Differential Polysome Association in Rice1[W][OA] , 2012, Plant Physiology.

[2]  Jukon Kim,et al.  Analysis of the APX, PGD1 and R1G1B constitutive gene promoters in various organs over three homozygous generations of transgenic rice plants , 2012, Planta.

[3]  Jukon Kim,et al.  Use of animal viral internal ribosome entry site sequence makes multiple truncated transcripts without mediating polycistronic expression in rice , 2011 .

[4]  R. Strasser,et al.  JIP analysis on rice (Oryza sativa cv Nipponbare) grown under limited nitrogen conditions , 2011 .

[5]  M. Crespi,et al.  Transcriptional and post-transcriptional regulation of a NAC1 transcription factor in Medicago truncatula roots. , 2011, The New phytologist.

[6]  J. Drenth,et al.  Overexpression of TaNAC69 leads to enhanced transcript levels of stress up-regulated genes and dehydration tolerance in bread wheat. , 2011, Molecular plant.

[7]  S. Balzergue,et al.  Characterization of a cinnamoyl-CoA reductase 1 (CCR1) mutant in maize: effects on lignification, fibre development, and global gene expression , 2011, Journal of experimental botany.

[8]  Jie Chen,et al.  Physiological mechanisms underlying OsNAC5-dependent tolerance of rice plants to abiotic stress , 2011, Planta.

[9]  R. Strasser,et al.  The use of JIP test to evaluate drought-tolerance of transgenic rice overexpressing OsNAC10 , 2011, Plant Biotechnology Reports.

[10]  K. Jung,et al.  Genetic and Molecular Insights into the Enhancement of Rice Yield Potential , 2011, Journal of Plant Biology.

[11]  Jinxing Lin,et al.  Development of Casparian strip in rice cultivars , 2011, Plant signaling & behavior.

[12]  Shoshi Kikuchi,et al.  Genome-wide analysis of NAC transcription factor family in rice. , 2010, Gene.

[13]  K. Shinozaki,et al.  The abiotic stress-responsive NAC-type transcription factor OsNAC5 regulates stress-inducible genes and stress tolerance in rice , 2010, Molecular Genetics and Genomics.

[14]  J. Lynch,et al.  Root cortical aerenchyma improves the drought tolerance of maize (Zea mays L.). , 2010, Plant, cell & environment.

[15]  R. E. Sharp,et al.  Complexity and coordination of root growth at low water potentials: recent advances from transcriptomic and proteomic analyses. , 2010, Plant, cell & environment.

[16]  Jukon Kim,et al.  Root-Specific Expression of OsNAC10 Improves Drought Tolerance and Grain Yield in Rice under Field Drought Conditions1[W][OA] , 2010, Plant Physiology.

[17]  Hui Peng,et al.  A NAC transcription factor gene of Chickpea (Cicer arietinum), CarNAC3, is involved in drought stress response and various developmental processes. , 2009, Journal of plant physiology.

[18]  M. Grusak,et al.  Identification of up-regulated genes in flag leaves during rice grain filling and characterization of OsNAC5, a new ABA-dependent transcription factor , 2009, Planta.

[19]  Zhi-hong Xu,et al.  Arabidopsis GLP4 is localized to the Golgi and binds auxin in vitro. , 2009, Acta biochimica et biophysica Sinica.

[20]  H. K. Park,et al.  Overexpression of the Transcription Factor AP37 in Rice Improves Grain Yield under Drought Conditions1[W][OA] , 2009, Plant Physiology.

[21]  Clayton T Larue,et al.  A microRNA-transcription factor module regulates lateral organ size and patterning in Arabidopsis. , 2009, The Plant journal : for cell and molecular biology.

[22]  B. Han,et al.  Overexpression of a NAC transcription factor enhances rice drought and salt tolerance. , 2009, Biochemical and biophysical research communications.

[23]  I. C. Lee,et al.  Trifurcate Feed-Forward Regulation of Age-Dependent Cell Death Involving miR164 in Arabidopsis , 2009, Science.

[24]  Jukon Kim,et al.  Rice NAC proteins act as homodimers and heterodimers , 2009, Plant Biotechnology Reports.

[25]  D. Llewellyn,et al.  The Low-Oxygen-Induced NAC Domain Transcription Factor ANAC102 Affects Viability of Arabidopsis Seeds following Low-Oxygen Treatment1[W][OA] , 2009, Plant Physiology.

[26]  Jukon Kim,et al.  Arabidopsis cyclin D2 expressed in rice forms a functional cyclin-dependent kinase complex that enhances seedling growth , 2008, Plant Biotechnology Reports.

[27]  T. Demura,et al.  Vascular-related NAC-DOMAIN7 is involved in the differentiation of all types of xylem vessels in Arabidopsis roots and shoots. , 2008, The Plant journal : for cell and molecular biology.

[28]  K. Theres,et al.  Interplay of miR164, CUP-SHAPED COTYLEDON genes and LATERAL SUPPRESSOR controls axillary meristem formation in Arabidopsis thaliana. , 2008, The Plant journal : for cell and molecular biology.

[29]  K. Shinozaki,et al.  Drought Induction of Arabidopsis 9-cis-Epoxycarotenoid Dioxygenase Occurs in Vascular Parenchyma Cells1[W][OA] , 2008, Plant Physiology.

[30]  L. Xiong,et al.  Characterization of transcription factor gene SNAC2 conferring cold and salt tolerance in rice , 2008, Plant Molecular Biology.

[31]  S. Song,et al.  Expression of barley HvCBF4 enhances tolerance to abiotic stress in transgenic rice. , 2007, Plant biotechnology journal.

[32]  K. Shinozaki,et al.  Functional analysis of a NAC-type transcription factor OsNAC6 involved in abiotic and biotic stress-responsive gene expression in rice. , 2007, The Plant journal : for cell and molecular biology.

[33]  Lizhong Xiong,et al.  Over-expression of a LEA gene in rice improves drought resistance under the field conditions , 2007, Theoretical and Applied Genetics.

[34]  J. Kehr,et al.  Long distance transport and movement of RNA through the phloem. , 2007, Journal of experimental botany.

[35]  C. Laloi,et al.  PDX1 is essential for vitamin B6 biosynthesis, development and stress tolerance in Arabidopsis. , 2006, The Plant journal : for cell and molecular biology.

[36]  L. Xiong,et al.  Overexpressing a NAM, ATAF, and CUC (NAC) transcription factor enhances drought resistance and salt tolerance in rice , 2006, Proceedings of the National Academy of Sciences.

[37]  R. Bressan,et al.  Osmogenetics: Aristotle to Arabidopsis , 2006, The Plant Cell Online.

[38]  Kazuo Shinozaki,et al.  Transcriptional regulatory networks in cellular responses and tolerance to dehydration and cold stresses. , 2006, Annual review of plant biology.

[39]  R. Yamamoto,et al.  Progressive Inhibition by Water Deficit of Cell Wall Extensibility and Growth along the Elongation Zone of Maize Roots Is Related to Increased Lignin Metabolism and Progressive Stelar Accumulation of Wall Phenolics1 , 2005, Plant Physiology.

[40]  P. McCourt,et al.  Molecular tailoring of farnesylation for plant drought tolerance and yield protection. , 2005, The Plant journal : for cell and molecular biology.

[41]  S. Kim,et al.  Arabidopsis CBF3/DREB1A and ABF3 in Transgenic Rice Increased Tolerance to Abiotic Stress without Stunting Growth1[w] , 2005, Plant Physiology.

[42]  Scott A. Givan,et al.  ASRP: the Arabidopsis Small RNA Project Database , 2004, Nucleic Acids Res..

[43]  E. Bray.  Genes commonly regulated by water-deficit stress in Arabidopsis thaliana. , 2004, Journal of experimental botany.

[44]  Kazuo Shinozaki,et al.  Isolation and Functional Analysis of Arabidopsis Stress-Inducible NAC Transcription Factors That Bind to a Drought-Responsive cis-Element in the early responsive to dehydration stress 1 Promoterw⃞ , 2004, The Plant Cell Online.

[45]  Kazuo Shinozaki,et al.  A dehydration-induced NAC protein, RD26, is involved in a novel ABA-dependent stress-signaling pathway. , 2004, The Plant journal : for cell and molecular biology.

[46]  K. Skriver,et al.  Structure of the conserved domain of ANAC, a member of the NAC family of transcription factors , 2004, EMBO reports.

[47]  Jianping Lu,et al.  CUPULIFORMIS establishes lateral organ boundaries in Antirrhinum , 2004, Development.

[48]  K. Shinozaki,et al.  Monitoring Expression Profiles of Rice Genes under Cold, Drought, and High-Salinity Stresses and Abscisic Acid Application Using cDNA Microarray and RNA Gel-Blot Analyses1[w] , 2003, Plant Physiology.

[49]  D. Hegedus,et al.  Molecular characterization of Brassicanapus NAC domain transcriptional activators induced in response to biotic and abiotic stress , 2003, Plant Molecular Biology.

[50]  J. Kawai,et al.  Collection, Mapping, and Annotation of Over 28,000 cDNA Clones from japonica Rice , 2003, Science.

[51]  S. D. de Vries,et al.  The CUP-SHAPED COTYLEDON3 Gene Is Required for Boundary and Shoot Meristem Formation in Arabidopsis Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.012203. , 2003, The Plant Cell Online.

[52]  M. K. Jensen,et al.  Interactions between plant RING-H2 and plant-specific NAC (NAM/ATAF1/2/CUC2) proteins: RING-H2 molecular specificity and cellular localization. , 2003, The Biochemical journal.

[53]  Y. Barrière,et al.  Down-regulation of the AtCCR1 gene in Arabidopsis thaliana: effects on phenotype, lignins and cell wall degradability , 2003, Planta.

[54]  S. Song,et al.  Expression of a Bifunctional Fusion of the Escherichia coli Genes for Trehalose-6-Phosphate Synthase and Trehalose-6-Phosphate Phosphatase in Transgenic Rice Plants Increases Trehalose Accumulation and Abiotic Stress Tolerance without Stunting Growth1 , 2003, Plant Physiology.

[55]  T. G. Owens,et al.  Trehalose accumulation in rice plants confers high tolerance levels to different abiotic stresses , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[56]  K. Akiyama,et al.  Monitoring the expression profiles of 7000 Arabidopsis genes under drought, cold and high-salinity stresses using a full-length cDNA microarray. , 2002, The Plant journal : for cell and molecular biology.

[57]  M. Thomashow,et al.  Arabidopsis Transcriptome Profiling Indicates That Multiple Regulatory Pathways Are Activated during Cold Acclimation in Addition to the CBF Cold Response Pathway Online version contains Web-only data. Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1 , 2002, The Plant Cell Online.

[58]  M. Oesterheld,et al.  Effects of Flooding and Drought on the Anatomy of Paspalum dilatatum , 2001 .

[59]  K. Shinozaki,et al.  Regulation of drought tolerance by gene manipulation of 9-cis-epoxycarotenoid dioxygenase, a key enzyme in abscisic acid biosynthesis in Arabidopsis. , 2001, The Plant journal : for cell and molecular biology.

[60]  A. R. Ennos,et al.  Cloning and characterization of irregular xylem4 (irx4): a severely lignin-deficient mutant of Arabidopsis. , 2001, The Plant journal : for cell and molecular biology.

[61]  R. R. Samaha,et al.  Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes. , 2000, Science.

[62]  N. Chua,et al.  Arabidopsis NAC1 transduces auxin signal downstream of TIR1 to promote lateral root development. , 2000, Genes & development.

[63]  Jukon Kim,et al.  Subcellular targeting of green fluorescent protein to plastids in transgenic rice plants provides a high-level expression system , 1999, Molecular Breeding.

[64]  E. Meyerowitz,et al.  A Homolog of NO APICAL MERISTEM Is an Immediate Target of the Floral Homeotic Genes APETALA3/PISTILLATA , 1998, Cell.

[65]  D. McCarty,et al.  Genetic control of abscisic acid biosynthesis in maize. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[66]  J. Medford,et al.  Modified MERI5 expression alters cell expansion in transgenic Arabidopsis plants , 1997 .

[67]  H Fujisawa,et al.  Genes involved in organ separation in Arabidopsis: an analysis of the cup-shaped cotyledon mutant. , 1997, The Plant cell.

[68]  P. Horton,et al.  REGULATION OF LIGHT HARVESTING IN GREEN PLANTS. , 1996, Annual review of plant physiology and plant molecular biology.

[69]  J. Mol,et al.  The No Apical Meristem Gene of Petunia Is Required for Pattern Formation in Embryos and Flowers and Is Expressed at Meristem and Primordia Boundaries , 1996, Cell.

[70]  T. Ho,et al.  Expression of a Late Embryogenesis Abundant Protein Gene, HVA1, from Barley Confers Tolerance to Water Deficit and Salt Stress in Transgenic Rice , 1996, Plant physiology.

[71]  T. Komari,et al.  Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. , 1994, The Plant journal : for cell and molecular biology.

[72]  E. Wurtele,et al.  An mRNA Putatively Coding for an O-Methyltransferase Accumulates Preferentially in Maize Roots and Is Located Predominantly in the Region of the Endodermis , 1993, Plant physiology.

[73]  L. Hensgens,et al.  The promoter of the rice gene GOS2 is active in various different monocot tissues and binds rice nuclear factor ASF-1. , 1992, The Plant journal : for cell and molecular biology.

[74]  Keith T. Ingram,et al.  Root Xylem Influence on the Water Relations and Drought Resistance of Rice , 1992 .

[75]  S. K. De Datta,et al.  Spikelet Sterility and Flowering Response of Rice to Water Stress at Anthesis , 1989 .

[76]  R. T. Cruz,et al.  Dryland Rice Response to an Irrigation Gradient at Flowering Stage1 , 1984 .

[77]  J. O’toole,et al.  Role of Panicle Exsertion in Water Stress Induced Sterility 1 , 1983 .

[78]  Yutaka Sato,et al.  Genome-wide transcriptome dissection of the rice root system: implications for developmental and physiological functions. , 2012, The Plant journal : for cell and molecular biology.

[79]  Jukon Kim,et al.  Accumulation of trehalose increases soluble sugar contents in rice plants conferring tolerance to drought and salt stress , 2011, Plant Biotechnology Reports.

[80]  Xi Chen,et al.  Evaluation of seven function-known candidate genes for their effects on improving drought resistance of transgenic rice under field conditions. , 2009, Molecular plant.

[81]  K. Shinozaki,et al.  Functional analysis of rice DREB1/CBF-type transcription factors involved in cold-responsive gene expression in transgenic rice. , 2006, Plant & cell physiology.

[82]  Yong Xu,et al.  Characterization of a rice gene family encoding root-specific proteins , 2004, Plant Molecular Biology.

[83]  Shoshi Kikuchi,et al.  Comprehensive analysis of NAC family genes in Oryza sativa and Arabidopsis thaliana. , 2003, DNA research : an international journal for rapid publication of reports on genes and genomes.

[84]  Heather Knight.  Calcium signaling during abiotic stress in plants. , 2000, International review of cytology.

[85]  Paul G. Falkowski,et al.  Photoinhibition of Photosynthesis in Nature , 1994 .