In Vivo Interference with AtTCP20 Function Induces Severe Plant Growth Alterations and Deregulates the Expression of Many Genes Important for Development[C][W]

AtTCP20 is a transcription factor belonging to the Arabidopsis (Arabidopsis thaliana) TCP-P subfamily, characterized by its capacity to bind to site II motifs (TGGGCY). Our aim was to understand the role of AtTCP20 in plant development. The expression pattern of a translational fusion of PromTCP20:CDS20∷GUS∷GFP suggested a function for AtTCP20 in several plant organs and stages of development. The role of AtTCP20 was challenged in planta by inducing expression of AtTCP20 proteins fused with either a transcriptional activator domain (VP16) or a repressor domain (EAR). Expression of both modified proteins led to severe developmental phenotypes. In-depth analysis suggested that AtTCP20 may participate in the regulation of cell expansion, cell division, and cell differentiation. Gene expression profiling in roots and hypocotyls revealed that 252 genes were down-regulated in both organs after induction of the AtTCP20∷EAR repressor gene. Site II motifs (TGGGCY) were underrepresented in their promoters. Conversely, GG(A/T)CCC sequences related to binding sites identified for TCP proteins in rice (Oryza sativa) were overrepresented, and a TCP20 fusion protein was shown to bind to these sequences in vitro. Gene ontology indicated that many targeted genes were involved in cell wall biogenesis and modification during expansion and also encoded numerous transcription factors controlling plant development. Our results are consistent with the previous proposal that AtTCP20 is involved in cell division and growth coordination. Moreover, they further suggest that AtTCP20 also contributes to cell expansion control and indicate a different involvement of this protein in plant morphogenesis depending on the organ and the developmental stage.

[1]  P. Benfey,et al.  Root development in Arabidopsis: four mutants with dramatically altered root morphogenesis. , 1993, Development.

[2]  S. Rhee,et al.  Functional Annotation of the Arabidopsis Genome Using Controlled Vocabularies1 , 2004, Plant Physiology.

[3]  A. Moorman,et al.  Assumption-free analysis of quantitative real-time polymerase chain reaction (PCR) data , 2003, Neuroscience Letters.

[4]  B Lescure,et al.  Plant interstitial telomere motifs participate in the control of gene expression in root meristems. , 1999, The Plant journal : for cell and molecular biology.

[5]  J. Ecker,et al.  Type-A Arabidopsis Response Regulators Are Partially Redundant Negative Regulators of Cytokinin Signaling Online version contains Web-only data. , 2004, The Plant Cell Online.

[6]  Naama Barkai,et al.  The design of transcription-factor binding sites is affected by combinatorial regulation , 2005, Genome Biology.

[7]  A. Bacic,et al.  The Fasciclin-Like Arabinogalactan Proteins of Arabidopsis. A Multigene Family of Putative Cell Adhesion Molecules1 , 2003, Plant Physiology.

[8]  Gordon K. Smyth,et al.  limma: Linear Models for Microarray Data , 2005 .

[9]  F. Skoog,et al.  A revised medium for rapid growth and bio assays with tobacco tissue cultures , 1962 .

[10]  J. Schiefelbein,et al.  The ROOT HAIR DEFECTIVE3 gene encodes an evolutionarily conserved protein with GTP-binding motifs and is required for regulated cell enlargement in Arabidopsis. , 1997, Genes & development.

[11]  P. Masson,et al.  Loss-of-Function Mutations of ROOT HAIR DEFECTIVE3 Suppress Root Waving, Skewing, and Epidermal Cell File Rotation in Arabidopsis1 , 2005, Plant Physiology.

[12]  Tomotsugu Koyama,et al.  TCP Transcription Factors Control the Morphology of Shoot Lateral Organs via Negative Regulation of the Expression of Boundary-Specific Genes in Arabidopsis[W][OA] , 2006, The Plant Cell Online.

[13]  Rafael A Irizarry,et al.  Exploration, normalization, and summaries of high density oligonucleotide array probe level data. , 2003, Biostatistics.

[14]  F. Parcy,et al.  Analysis of an activated ABI5 allele using a new selection method for transgenic Arabidopsis seeds , 2004, FEBS letters.

[15]  H. Fukuda,et al.  HD-zip III homeobox genes that include a novel member, ZeHB-13 (Zinnia)/ATHB-15 (Arabidopsis), are involved in procambium and xylem cell differentiation. , 2003, Plant & cell physiology.

[16]  B. Sundberg,et al.  hca: an Arabidopsis mutant exhibiting unusual cambial activity and altered vascular patterning. , 2005, The Plant journal : for cell and molecular biology.

[17]  N. Chua,et al.  KORRIGAN, an Arabidopsis Endo-1,4-β-Glucanase, Localizes to the Cell Plate by Polarized Targeting and Is Essential for Cytokinesis , 2000, Plant Cell.

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

[19]  S. Clough,et al.  Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. , 1998, The Plant journal : for cell and molecular biology.

[20]  Y. Kamiya,et al.  Identification of cis-Elements That Regulate Gene Expression during Initiation of Axillary Bud Outgrowth in Arabidopsis[w] , 2005, Plant Physiology.

[21]  D. Baulcombe,et al.  Suppression of gene silencing: a general strategy used by diverse DNA and RNA viruses of plants. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[22]  D. Tremousaygue,et al.  TCP Transcription Factors Predate the Emergence of Land Plants , 2007, Journal of Molecular Evolution.

[23]  S. Tiwari,et al.  Aux/IAA Proteins Contain a Potent Transcriptional Repression Domain , 2004, The Plant Cell Online.

[24]  G. Crooks,et al.  WebLogo: A sequence logo generator, Genome Research, , 2004 .

[25]  P. Zimmermann,et al.  GENEVESTIGATOR. Arabidopsis Microarray Database and Analysis Toolbox1[w] , 2004, Plant Physiology.

[26]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[27]  Yuko Ohashi,et al.  DNA binding and dimerization specificity and potential targets for the TCP protein family. , 2002, The Plant journal : for cell and molecular biology.

[28]  L. Garnier,et al.  Internal telomeric repeats and 'TCP domain' protein-binding sites co-operate to regulate gene expression in Arabidopsis thaliana cycling cells. , 2003, The Plant journal : for cell and molecular biology.

[29]  M. Pfaffl,et al.  A new mathematical model for relative quantification in real-time RT-PCR. , 2001, Nucleic acids research.

[30]  E. Coen,et al.  Origin of floral asymmetry in Antirrhinum , 1996, Nature.

[31]  Laurie G. Smith,et al.  Spatial control of cell expansion by the plant cytoskeleton. , 2005, Annual review of cell and developmental biology.

[32]  S. Kosugi,et al.  Two of three promoter elements identified in a rice gene for proliferating cell nuclear antigen are essential for meristematic tissue-specific expression. , 1995, The Plant journal : for cell and molecular biology.

[33]  D. Weigel,et al.  A genetic framework for floral patterning , 1998, Nature.

[34]  K. Hiratsu,et al.  A chimeric AtMYB23 repressor induces hairy roots, elongation of leaves and stems, and inhibition of the deposition of mucilage on seed coats in Arabidopsis. , 2005, Plant & cell physiology.

[35]  Jian-Kang Zhu,et al.  The Arabidopsis SOS5 Locus Encodes a Putative Cell Surface Adhesion Protein and Is Required for Normal Cell Expansion Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.007872. , 2003, The Plant Cell Online.

[36]  N. Chua,et al.  Chemical-inducible systems for regulated expression of plant genes. , 2000, Current opinion in biotechnology.

[37]  D. Gonzalez,et al.  Overrepresentation of Elements Recognized by TCP-Domain Transcription Factors in the Upstream Regions of Nuclear Genes Encoding Components of the Mitochondrial Oxidative Phosphorylation Machinery1[W] , 2006, Plant Physiology.

[38]  P. Benfey,et al.  Conditional root expansion mutants of Arabidopsis. , 1995, Development.

[39]  Peter Doerner,et al.  Arabidopsis TCP20 links regulation of growth and cell division control pathways. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[40]  Jin-Young Park,et al.  Genome-wide expression profiling of ARABIDOPSIS RESPONSE REGULATOR 7(ARR7) overexpression in cytokinin response , 2007, Molecular Genetics and Genomics.

[41]  S. Tabata,et al.  RNA Interference of the Arabidopsis Putative Transcription Factor TCP16 Gene Results in Abortion of Early Pollen Development , 2006, Plant Molecular Biology.

[42]  J. Doebley,et al.  The evolution of apical dominance in maize , 1997, Nature.

[43]  G. Crooks,et al.  WebLogo: a sequence logo generator. , 2004, Genome research.

[44]  S. Kosugi,et al.  PCF1 and PCF2 specifically bind to cis elements in the rice proliferating cell nuclear antigen gene. , 1997, The Plant cell.

[45]  Makoto Matsuoka,et al.  The OsTB1 gene negatively regulates lateral branching in rice. , 2003, The Plant journal : for cell and molecular biology.

[46]  K. Hiratsu,et al.  Dominant repression of target genes by chimeric repressors that include the EAR motif, a repression domain, in Arabidopsis. , 2003, The Plant journal : for cell and molecular biology.

[47]  Christopher D Town,et al.  Development and evaluation of an Arabidopsis whole genome Affymetrix probe array. , 2004, The Plant journal : for cell and molecular biology.

[48]  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.

[49]  Y. Benjamini,et al.  Controlling the false discovery rate: a practical and powerful approach to multiple testing , 1995 .

[50]  Ana I. Caño-Delgado,et al.  The eli1 mutation reveals a link between cell expansion and secondary cell wall formation in Arabidopsis thaliana. , 2000, Development.

[51]  Jennifer Hayes Clark,et al.  Control of Organ Asymmetry in Flowers of Antirrhinum , 1999, Cell.

[52]  Yan Wang,et al.  The Arabidopsis homeobox gene, ATHB16, regulates leaf development and the sensitivity to photoperiod in Arabidopsis. , 2003, Developmental biology.

[53]  Maureen C. McCann,et al.  Cell-wall structure and anisotropy in procuste, a cellulose synthase mutant of Arabidopsis thaliana , 2006, Planta.

[54]  T. Baskin,et al.  Temperature-sensitive alleles of RSW2 link the KORRIGAN endo-1,4-beta-glucanase to cellulose synthesis and cytokinesis in Arabidopsis. , 2001, Plant physiology.

[55]  Renze Heidstra,et al.  Cytokinins Determine Arabidopsis Root-Meristem Size by Controlling Cell Differentiation , 2007, Current Biology.

[56]  J. Hanson,et al.  Homeodomain Leucine Zipper Class I Genes in Arabidopsis. Expression Patterns and Phylogenetic Relationships1[w] , 2005, Plant Physiology.

[57]  G. Coupland,et al.  A Dissociation insertion causes a semidominant mutation that increases expression of TINY, an Arabidopsis gene related to APETALA2. , 1996, The Plant cell.

[58]  V. Gomord,et al.  Cloning and sequence analysis of laccase-encoding cDNA clones from tobacco. , 1996, Gene.

[59]  E. Coen,et al.  Genetic Control of Surface Curvature , 2003, Science.

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

[61]  Thorsten Hamann,et al.  Developmental specificity of auxin response by pairs of ARF and Aux/IAA transcriptional regulators , 2005, The EMBO journal.

[62]  Kazuo Shinozaki,et al.  AREB1 Is a Transcription Activator of Novel ABRE-Dependent ABA Signaling That Enhances Drought Stress Tolerance in Arabidopsis[W][OA] , 2005, The Plant Cell Online.

[63]  Jean YH Yang,et al.  Bioconductor: open software development for computational biology and bioinformatics , 2004, Genome Biology.

[64]  Lisa Newman,et al.  Involvement of the R2R3-MYB, AtMYB61, in the ectopic lignification and dark-photomorphogenic components of the det3 mutant phenotype. , 2004, The Plant journal : for cell and molecular biology.

[65]  Y. van de Peer,et al.  Identification of novel regulatory modules in dicotyledonous plants using expression data and comparative genomics , 2006, Genome Biology.

[66]  Broome,et al.  Literature cited , 1924, A Guide to the Carnivores of Central America.

[67]  J. Kieber,et al.  Cytokinin signaling. , 2005, Current opinion in plant biology.

[68]  K. Nishitani,et al.  A principal role for AtXTH18 in Arabidopsis thaliana root growth: a functional analysis using RNAi plants , 2006, Journal of Plant Research.

[69]  J. Doebley,et al.  teosinte branched1 and the origin of maize: evidence for epistasis and the evolution of dominance. , 1995, Genetics.

[70]  E. Coen,et al.  The TCP domain: a motif found in proteins regulating plant growth and development. , 1999, The Plant journal : for cell and molecular biology.