FLOWERING BHLH transcriptional activators control expression of the photoperiodic flowering regulator CONSTANS in Arabidopsis

Many plants monitor day-length changes throughout the year and use the information to precisely regulate the timing of seasonal flowering for maximum reproductive success. In Arabidopsis thaliana, transcriptional regulation of the CONSTANS (CO) gene and posttranslational regulation of CO protein are crucial mechanisms for proper day-length measurement in photoperiodic flowering. Currently, the CYCLING DOF FACTOR proteins are the only transcription factors known to directly regulate CO gene expression, and the mechanisms that directly activate CO transcription have remained unknown. Here we report the identification of four CO transcriptional activators, named FLOWERING BHLH 1 (FBH1), FBH2, FBH3, and FBH4. All FBH proteins are related basic helix–loop–helix-type transcription factors that preferentially bind to the E-box cis-elements in the CO promoter. Overexpression of all FBH genes drastically elevated CO levels and caused early flowering regardless of photoperiod, whereas CO levels were reduced in the fbh quadruple mutants. In addition, FBH1 is expressed in the vascular tissue and bound near the transcription start site of the CO promoter in vivo. Furthermore, FBH homologs in poplar and rice induced CO expression in Arabidopsis. These results indicate that FBH proteins positively regulate CO transcription for photoperiodic flowering and that this mechanism may be conserved in diverse plant species. Our results suggest that the diurnal CO expression pattern is generated by a concert of redundant functions of positive and negative transcriptional regulators.

[1]  S. Hendricks,et al.  Photoperiodism in Plants. , 1960, Science.

[2]  Daphne Vince-Prue,et al.  Photoperiodism in Plants , 1975 .

[3]  L. Sieburth,et al.  Molecular dissection of the AGAMOUS control region shows that cis elements for spatial regulation are located intragenically. , 1997, The Plant cell.

[4]  D. P. King,et al.  Role of the CLOCK protein in the mammalian circadian mechanism. , 1998, Science.

[5]  R. Amasino,et al.  FLOWERING LOCUS C Encodes a Novel MADS Domain Protein That Acts as a Repressor of Flowering , 1999, Plant Cell.

[6]  P. Hardin,et al.  Interlocked feedback loops within the Drosophila circadian oscillator. , 1999, Science.

[7]  G. Coupland,et al.  GIGANTEA: a circadian clock‐controlled gene that regulates photoperiodic flowering in Arabidopsis and encodes a protein with several possible membrane‐spanning domains , 1999, The EMBO journal.

[8]  N. Chua,et al.  Technical advance: An estrogen receptor-based transactivator XVE mediates highly inducible gene expression in transgenic plants. , 2000, The Plant journal : for cell and molecular biology.

[9]  Claire Périlleux,et al.  Mutagenesis of Plants Overexpressing CONSTANS Demonstrates Novel Interactions among Arabidopsis Flowering-Time Genes , 2000, Plant Cell.

[10]  Hitoshi Onouchi,et al.  CONSTANS mediates between the circadian clock and the control of flowering in Arabidopsis , 2001, Nature.

[11]  S. Kay,et al.  Living by the calendar: how plants know when to flower , 2003, Nature Reviews Molecular Cell Biology.

[12]  Koji Goto,et al.  TERMINAL FLOWER2, an Arabidopsis Homolog of HETEROCHROMATIN PROTEIN1, Counteracts the Activation of FLOWERING LOCUS T by CONSTANS in the Vascular Tissues of Leaves to Regulate Flowering Time Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.016 , 2003, The Plant Cell Online.

[13]  S. Kay,et al.  FKF1 is essential for photoperiodic-specific light signalling in Arabidopsis , 2003, Nature.

[14]  E. Huq,et al.  The Arabidopsis Basic/Helix-Loop-Helix Transcription Factor Family Online version contains Web-only data. Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.013839. , 2003, The Plant Cell Online.

[15]  Shelley Hepworth,et al.  CONSTANS acts in the phloem to regulate a systemic signal that induces photoperiodic flowering of Arabidopsis , 2004, Development.

[16]  D. Ravenscroft,et al.  Photoreceptor Regulation of CONSTANS Protein in Photoperiodic Flowering , 2004, Science.

[17]  K. Goto,et al.  FD, a bZIP Protein Mediating Signals from the Floral Pathway Integrator FT at the Shoot Apex , 2005, Science.

[18]  G. Coupland,et al.  Distinct Roles of GIGANTEA in Promoting Flowering and Regulating Circadian Rhythms in Arabidopsis , 2005, The Plant Cell Online.

[19]  Wolfgang Busch,et al.  Integration of Spatial and Temporal Information During Floral Induction in Arabidopsis , 2005, Science.

[20]  S. Kay,et al.  FKF1 F-Box Protein Mediates Cyclic Degradation of a Repressor of CONSTANS in Arabidopsis , 2005, Science.

[21]  Christian A. Grove,et al.  A Gene-Centered C. elegans Protein-DNA Interaction Network , 2006, Cell.

[22]  Yuhong Yang,et al.  CKI and CKII mediate the FREQUENCY-dependent phosphorylation of the WHITE COLLAR complex to close the Neurospora circadian negative feedback loop. , 2006, Genes & development.

[23]  Mingjie Chen,et al.  RFI2, a RING-domain zinc finger protein, negatively regulates CONSTANS expression and photoperiodic flowering. , 2006, The Plant journal : for cell and molecular biology.

[24]  P. Hardin,et al.  PER-dependent rhythms in CLK phosphorylation and E-box binding regulate circadian transcription. , 2006, Genes & development.

[25]  Stefan Jansson,et al.  CO/FT Regulatory Module Controls Timing of Flowering and Seasonal Growth Cessation in Trees , 2006, Science.

[26]  Fabio Fornara,et al.  FT Protein Movement Contributes to Long-Distance Signaling in Floral Induction of Arabidopsis , 2007, Science.

[27]  S. Kim,et al.  Control of Flowering Time and Cold Response by a NAC-Domain Protein in Arabidopsis , 2007, PloS one.

[28]  Dmitri A. Nusinow,et al.  FKF1 and GIGANTEA Complex Formation Is Required for Day-Length Measurement in Arabidopsis , 2007, Science.

[29]  D. Weigel,et al.  Move on up, it's time for change--mobile signals controlling photoperiod-dependent flowering. , 2007, Genes & development.

[30]  Young Hun Song,et al.  Isolation of CONSTANS as a TGA4/OBF4 interacting protein. , 2008, Molecules and cells.

[31]  Y. Wang,et al.  Two New Clock Proteins, LWD1 and LWD2, Regulate Arabidopsis Photoperiodic Flowering1[W][OA] , 2008, Plant Physiology.

[32]  H. Nam,et al.  FIONA1 Is Essential for Regulating Period Length in the Arabidopsis Circadian Clock[W] , 2008, The Plant Cell Online.

[33]  Ghislain Breton,et al.  A Functional Genomics Approach Reveals CHE as a Component of the Arabidopsis Circadian Clock , 2009, Science.

[34]  G. Coupland,et al.  Arabidopsis DOF transcription factors act redundantly to reduce CONSTANS expression and are essential for a photoperiodic flowering response. , 2009, Developmental cell.

[35]  Y. Fukada,et al.  Roles of CLOCK Phosphorylation in Suppression of E-Box-Dependent Transcription , 2009, Molecular and Cellular Biology.

[36]  J. Casaretto,et al.  VvCO and VvCOL1, two CONSTANS homologous genes, are regulated during flower induction and dormancy in grapevine buds , 2009, Plant Cell Reports.

[37]  D. Robertson,et al.  Genome-Wide Classification and Evolutionary Analysis of the bHLH Family of Transcription Factors in Arabidopsis, Poplar, Rice, Moss, and Algae1[W] , 2010, Plant Physiology.

[38]  R. Tóth,et al.  Plant development goes like clockwork. , 2010, Trends in genetics : TIG.

[39]  Young Hun Song,et al.  Similarities in the circadian clock and photoperiodism in plants. , 2010, Current opinion in plant biology.

[40]  T. Imaizumi,et al.  Arabidopsis circadian clock and photoperiodism: time to think about location. , 2010, Current opinion in plant biology.

[41]  R. Amasino.  Seasonal and developmental timing of flowering. , 2010, The Plant journal : for cell and molecular biology.

[42]  L. Dolan,et al.  Origin and Diversification of Basic-Helix-Loop-Helix Proteins in Plants , 2009, Molecular biology and evolution.