The Arabidopsis RGA Gene Encodes a Transcriptional Regulator Repressing the Gibberellin Signal Transduction Pathway

The recessive rga mutation is able to partially suppress phenotypic defects of the Arabidopsis gibberellin (GA) biosynthetic mutant ga1-3. Defects in stem elongation, flowering time, and leaf abaxial trichome initiation are suppressed by rga. This indicates that RGA is a negative regulator of the GA signal transduction pathway. We have identified 10 additional alleles of rga from a fast-neutron mutagenized ga1-3 population and used them to isolate the RGA gene by genomic subtraction. Our data suggest that RGA may be functioning as a transcriptional regulator. RGA was found to be a member of the VHIID regulatory family, which includes the radial root organizing gene SCARECROW and another GA signal transduction repressor, GAI. RGA and GAI proteins share a high degree of homology, but their N termini are more divergent. The presence of several structural features, including homopolymeric serine and threonine residues, a putative nuclear localization signal, leucine heptad repeats, and an LXXLL motif, indicates that the RGA protein may be a transcriptional regulator that represses the GA response. In support of the putative nuclear localization signal, we demonstrated that a transiently expressed green fluorescent protein–RGA fusion protein is localized to the nucleus in onion epidermal cells. Because the rga mutation abolished the high level of expression of the GA biosynthetic gene GA4 in the ga1-3 mutant background, we conclude that RGA may also play a role in controlling GA biosynthesis.

[1]  P. Benfey,et al.  The SCARECROW Gene Regulates an Asymmetric Cell Division That Is Essential for Generating the Radial Organization of the Arabidopsis Root , 1996, Cell.

[2]  M. Kanehisa,et al.  A knowledge base for predicting protein localization sites in eukaryotic cells , 1992, Genomics.

[3]  T. Sun,et al.  Developmental regulation of the gibberellin biosynthetic gene GA1 in Arabidopsis thaliana. , 1997, The Plant journal : for cell and molecular biology.

[4]  D. Prasher,et al.  Removal of a cryptic intron and subcellular localization of green fluorescent protein are required to mark transgenic Arabidopsis plants brightly. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[5]  M. Caboche,et al.  Sequence and characterization of two Arabidopsis thaliana cDNAs isolated by functional complementation of a yeast gln3 gdh1 mutant , 1997, FEBS letters.

[6]  M. H. Beale,et al.  Probing Gibberellin Receptors in the Avena fatua Aleurone , 1991 .

[7]  I. Hwang,et al.  Isolation of the Arabidopsis GA4 locus. , 1995, The Plant cell.

[8]  C. Somerville,et al.  Cellular differentiation regulated by gibberellin in the Arabidopsis thaliana pickle mutant. , 1997, Science.

[9]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[10]  J. Hanover,et al.  O-Linked GlcNAc Transferase Is a Conserved Nucleocytoplasmic Protein Containing Tetratricopeptide Repeats* , 1997, The Journal of Biological Chemistry.

[11]  Y. Kamiya,et al.  GIBBERELLIN BIOSYNTHESIS: Enzymes, Genes and Their Regulation. , 1997, Annual review of plant physiology and plant molecular biology.

[12]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[13]  A. Riggs,et al.  Genomic sequencing. , 1993, Methods in molecular biology.

[14]  P. McCourt,et al.  A Protein Farnesyl Transferase Involved in Abscisic Acid Signal Transduction in Arabidopsis , 1996, Science.

[15]  P. Hedden,et al.  Isolation and Expression of Three Gibberellin 20-Oxidase cDNA Clones from Arabidopsis , 1995, Plant physiology.

[16]  H. Goodman,et al.  Effects of ionizing radiation on a plant genome: analysis of two Arabidopsis transparent testa mutations. , 1992, The Plant cell.

[17]  N. Raikhel.  Nuclear targeting in plants. , 1992, Plant physiology.

[18]  Maarten Koornneef,et al.  A gibberellin insensitive mutant of Arabidopsis thaliana , 1985 .

[19]  T. Sun,et al.  The Arabidopsis GA1 locus encodes the cyclase ent-kaurene synthetase A of gibberellin biosynthesis. , 1994, The Plant cell.

[20]  H. Goodman,et al.  The genome of Arabidopsis thaliana. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

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

[22]  M. Koornneef,et al.  Genetic fine structure of the ga-1 locus in the higher plant Arabidopsis thaliana , 1983 .

[23]  J. Peng,et al.  The Arabidopsis GAI gene defines a signaling pathway that negatively regulates gibberellin responses. , 1997, Genes & development.

[24]  A. Peeters,et al.  The GA5 locus of Arabidopsis thaliana encodes a multifunctional gibberellin 20-oxidase: molecular cloning and functional expression. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[25]  R. N. Wilson,et al.  Phenotypic Suppression of the Gibberellin-Insensitive Mutant (gai) of Arabidopsis , 1995, Plant physiology.

[26]  P. J. Davies.  Plant hormones : physiology, biochemistry and molecular biology , 1995 .

[27]  S. Jacobsen,et al.  SPINDLY, a tetratricopeptide repeat protein involved in gibberellin signal transduction in Arabidopsis. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[28]  David M. Heery,et al.  A signature motif in transcriptional co-activators mediates binding to nuclear receptors , 1997, Nature.

[29]  J. Peng,et al.  Derivative Alleles of the Arabidopsis Gibberellin-Insensitive (gai) Mutation Confer a Wild-Type Phenotype. , 1993, The Plant cell.

[30]  M. Koornneef,et al.  EMS- and radiation-induced mutation frequencies at individual loci in Arabidopsis thaliana (L.) Heynh. , 1982, Mutation research.

[31]  H. Goodman,et al.  Cloning the Arabidopsis GA1 Locus by Genomic Subtraction. , 1992, The Plant cell.

[32]  G. Hart,et al.  Dynamic Glycosylation of Nuclear and Cytosolic Proteins , 1997, The Journal of Biological Chemistry.

[33]  C. Gasser,et al.  Cloning and characterization of chloroplast and cytosolic forms of cyclophilin from Arabidopsis thaliana. , 1994, The Journal of biological chemistry.

[34]  C. Lister,et al.  Recombinant inbred lines for mapping RFLP and phenotypic markers in Arabidopsis thaliana , 1993 .

[35]  S. Jacobsen,et al.  Mutations at the SPINDLY locus of Arabidopsis alter gibberellin signal transduction. , 1993, The Plant cell.

[36]  L. Cattivelli,et al.  Characterization of two barley genes that respond rapidly to dehydration stress , 1995 .

[37]  G. Storz,et al.  Analysis of fast neutron-generated mutants at the Arabidopsis thaliana HY4 locus. , 1996, The Plant journal : for cell and molecular biology.

[38]  S. Gilroy,et al.  Perception of Gibberellin and Abscisic Acid at the External Face of the Plasma Membrane of Barley (Hordeum vulgare L.) Aleurone Protoplasts , 1994, Plant physiology.

[39]  S. Swain,et al.  Genetic Analysis of Gibberellin Signal Transduction , 1996, Plant physiology.

[40]  N. Raikhel,et al.  Nuclear localization signal(s) required for nuclear targeting of the maize regulatory protein Opaque-2. , 1992, The Plant cell.

[41]  T. Sun,et al.  The new RGA locus encodes a negative regulator of gibberellin response in Arabidopsis thaliana. , 1997, Genetics.

[42]  F. Ausubel,et al.  Specialized binary vector for plant transformation: expression of the Arabidopsis thaliana AHAS gene in Nicotiana tabacum. , 1988, Nucleic acids research.

[43]  M. Koornneef,et al.  EMS- and relation-induced mutation frequencies at individual loci in Arabidopsis thaliana (L.) Heynh , 1982 .

[44]  Christopher K. Glass,et al.  The transcriptional co-activator p/CIP binds CBP and mediates nuclear-receptor function , 1997, Nature.