Florigen goes molecular: Seventy years of the hormonal theory of flowering regulation

As early as in 1936, the comprehensive studies of flowering led M.Kh. Chailakhyan to the concept of florigen, a hormonal floral stimulus, and let him establish several characteristics of this stimulus. These studies set up for many years the main avenues for research into the processes that control plant flowering, and the notion of florigen became universally accepted by scientists worldwide. The present-day evidence of genetic control of plant flowering supports the idea that florigen participates in floral signal transduction. The recent study of arabidopsis plants led the authors to conclusion that the immediate products of the gene FLOWERING LOCUS I, its mRNA and/or protein, move from an induced leaf into the shoot apex and evoke flowering therein.     

Molecular Genetics Returns to Basic Postulates of the Florigen Theory

The florigen theory, also known as the hormonal theory of flowering in plants, was put forward by M.Kh. Chailakhyan more than 65 years ago. This theory, reviewed here in its historical progress, gave an impetus to a number of modern genetic and molecular studies of flowering in plants. In turn, results of recent investigations restored to a new level Chailakhyan's ideas about florigen, the hormonal stimulus of flowering.     

Hormonal regulation of reproductive development in higher plants

The present paper deals with the hormonal regulation of reproductive development,i.e. flowering and sex manifestation. Representation of hormonal regulation of flowering is based on the concept of florigen as a two-component, complementary system of flowering hormones, which consists of gibberellins and anthesins. Data are presented on the effect of extracts of gibberellin- and anthesin-type substances from the leaves of flowering and vegetatively growing plants under non-inductive conditions of day length. Experiments with flowering of plants under the influence of graftings serve as a basis for considering the question about the common nature of one of the florigen components — anthesins, for various plant species. The mechanism controlling tuberization in both intact and grafted plants is based on the participation of all the components of the hormonal system and constitutes one of the most vivid manifestations of integration of all the organs in the whole organism.     

FTIP1 is an essential regulator required for florigen transport

The capacity to respond to day length, photoperiodism, is crucial for flowering plants to adapt to seasonal change. The photoperiodic control of flowering in plants is mediated by a long-distance mobile floral stimulus called florigen that moves from leaves to the shoot apex. Although the proteins encoded by FLOWERING LOCUS T (FT) in Arabidopsis and its orthologs in other plants are identified as the long-sought florigen, whether their transport is a simple diffusion process or under regulation remains elusive. Here we show that an endoplasmic reticulum (ER) membrane protein, FT-INTERACTING PROTEIN 1 (FTIP1), is an essential regulator required for FT protein transport in Arabidopsis. Loss of function of FTIP1 exhibits late flowering under long days, which is partly due to the compromised FT movement to the shoot apex. FTIP1 and FT share similar mRNA expression patterns and subcellular localization, and they interact specifically in phloem companion cells. FTIP1 is required for FT export from companion cells to sieve elements, thus affecting FT transport through the phloem to the SAM. Our results provide a mechanistic understanding of florigen transport, demonstrating that FT moves in a regulated manner and that FTIP1 mediates FT transport to induce flowering.     

ZCN8 encodes a potential orthologue of Arabidopsis FT florigen that integrates both endogenous and photoperiod flowering signals in maize

Higher plants use multiple perceptive measures to coordinate flowering time with environmental and endogenous cues. Physiological studies show that florigen is a mobile factor that transmits floral inductive signals from the leaf to the shoot apex. Arabidopsis FT protein is widely regarded as the archetype florigen found in diverse plant species, particularly in plants that use inductive photoperiods to flower. Recently, a large family of FT homologues in maize, the Zea CENTRORADIALIS (ZCN) genes, was described, suggesting that maize also contains FT-related proteins that act as a florigen. The product of one member of this large family, ZCN8, has several attributes that make it a good candidate as a maize florigen. Mechanisms underlying the floral transition in maize are less well understood than those of other species, partly because flowering in temperate maize is dependent largely on endogenous signals. The maize indeterminate1 (id1) gene is an important regulator of maize autonomous flowering that acts in leaves to mediate the transmission or production of florigenic signals. This study finds that id1 acts upstream of ZCN8 to control its expression, suggesting a possible new link to flowering in day-neutral maize. Moreover, in teosinte, a tropical progenitor of maize that requires short-day photoperiods to induce flowering, ZCN8 is highly up-regulated in leaves under inductive photoperiods. Finally, vascular-specific expression of ZCN8 in Arabidopsis complements the ft-1 mutation, demonstrating that leaf-specific expression of ZCN8 can induce flowering. These results suggest that ZCN8 may encode a florigen that integrates both endogenous and environmental signals in maize.     

Regulation of flowering plants of different biotypes

The patterns of control of flowering are analyzed in plants of different biotypes. The photoperiodic reaction of flowering taken as an example, the whole net of control is considered: from the environmental stimulus through its physiological transformation in the leaf in the corresponding hormonal impulse which, in turn, controls the realization of genetic programme and formation of generative organs in the stem buds. The photoperiodically neutral plants taken as an example, the patterns of age control of flowering are considered. In plants of different photoperiodic groups the synthesis of complementary components of florigen was shown to proceed either autonomously under the photoperiodic effect or under the inducing effect of definite photoperiods. The autonomous and inducible mechanisms of biosynthesis of the flowering hormones have a common base, the genetic system to which the environment sends its stimuli through the hormonal interactions. The interaction of hormonal and genetic developmental factors is considered, the evocation of flowering in the stem buds taken as an example.     

A florigenic effect of sucrose in Fuchsia hybrida is blocked by gibberellin-induced assimilate competition

The use of gas chromatography-mass spectrometry-selected ion monitoring along with a (13)C internal standard has allowed sensitive measurements of the sucrose (Suc) content of individual shoot apices of Fuchsia hybrida. With intact plants, as the photosynthetic irradiance increased, so did shoot apex Suc content, reaching saturation at about 500 micromol m(-2) s(-1). These same plants flowered at the higher irradiances, remaining vegetative in 10-h short days at an irradiance of 230 micromol m(-2) s(-1). The strong correlation (r = 0.93) in these studies between flowering and shoot apex Suc content indicates a role for Suc as a stimulus to flowering in this species. However, Suc is not the long-day (LD) "florigen" of F. hybrida because 2 to 4 LD given as a 14-h low-irradiance photoperiod extension (10-15 micromol m(-2) s(-1)) induced flowering but without increase in shoot apex Suc content. Flowering induced by either pathway, the LD- or the Suc-mediated one, was inhibited by applying gibberellin (GA) to the shoot tip. Such inhibition of flowering by GA, at least for the LD pathway, was associated with a reduced apex Suc content, enhanced elongation of subapical stem tissue, and a reduced import into the shoot apex of leaf-sourced assimilate. Thus, our findings show how GA inhibits flowering of F. hybrida and confirm the importance of nutrient diversion in regulating flowering.     

Universal florigenic signals triggered by FT homologues regulate growth and flowering cycles in perennial day-neutral tomato

The transition from vegetative to floral meristems in higher plants is programmed by the coincidence of internal and environmental signals. Classic grafting experiments have shown that leaves, in response to changing photoperiods, emit systemic signals, dubbed 'florigen', which induce flowering at the shoot apex. The florigen paradigm was conceived in photoperiod-sensitive plants: nevertheless it implies that although activated by different stimuli in different flowering systems, the signal is common to all plants. Tomato is a day-neutral, perennial plant, with sympodial and modular organization of its shoots and thus with reiterative regular vegetative/reproductive transitions. SINGLE FLOWER TRUSS a regulator of flowering-time and shoot architecture encodes the tomato orthologue of FT, a major flowering integrator gene in Arabidopsis. SFT generates graft-transmissible signals which complement the morphogenetic defects in sft plants, substitute for light dose stimulus in tomato and for contrasting day-length requirements in Arabidopsis and MARYLAND MAMMOTH tobacco. It is discussed how systemic signals initiated by SFT interact with the SELF PRUNING gene to regulate vegetative to reproductive (V/R) transitions in the context of two flowering systems, one for primary apices and the other for sympodial shoots.     

Control of water-use efficiency by florigen

A major issue in modern agriculture is water loss through stomata during photosynthetic carbon assimilation. In water-limited ecosystems, annual plants have strategies to synchronize their growth and reproduction to the availability of water. Some species or ecotypes of flowers are early to ensure that their life cycles are completed before the onset of late season terminal drought (“drought escape”). This accelerated flowering correlates with low water-use efficiency (WUE). The molecular players and physiological mechanisms involved in this coordination are not fully understood. We analyzed WUE using gravimetry, gas exchange, and carbon isotope discrimination in florigen deficient (sft mutant), wild-type (Micro-Tom), and florigen over-expressing (SFT-ox) tomato lines. Increased florigen expression led to accelerated flowering time and reduced WUE. The low WUE of SFT-ox was driven by higher stomatal conductance and thinner leaf blades. This florigen-driven effect on WUE appears be independent of abscisic acid (ABA). Our results open a new avenue to increase WUE in crops in an ABA-independent manner. Manipulation of florigen levels could allow us to produce crops with a life cycle synchronized to water availability.     

Florigen （Ⅱ）： It is a Mobile Protein

The true identity of florigen - the molecule（s） that migrates from leaves to apical meristem to initiate flowering - was notoriously elusive, having made it almost the ＂Bigfoot＂ of plant biology. There was never a lack of drama in the field of florigen study, and florigen researchers have once again experienced such a swing in the last two years. We wrote a minireview last year in this journal （Yu et al. 2006） to excitedly salute, among other discoveries, the notion that the flowering locus T （FT） mRNA might be the molecular form of a florigen. However, this hypothesis was challenged in a little less than two years after its initial proposition, and the original paper proposed that the FT mRNA hypothesis was retracted （Huang et al. 2005; Bohlenius et al. 2007）. Interestingly enough, the FT gene previously proposed to encode a florigen was never challenged. Rather, the FT protein, instead of the FT mRNA, is now believed to migrate from leaves to the apical meristem to promote floral initiation. In this update, we will share with our readers some entertaining stories concerning the recent studies of florigen in five different plant species. In addition to the published reports referenced inthis update, readers may also refer to our previous minireview and references therein for additional background information （Yu et al. 2006）.     

Florigen is involved in axillary bud development at multiple stages in Arabidopsis

The wide variety of plant architectures is largely based on diverse and flexible modes of axillary shoot development. In Arabidopsis, floral transition (flowering) stimulates axillary bud development. The mechanism that links flowering and axillary bud development is, however, largely unknown. We recently showed that FLOWERING LOCUS T (FT) protein, which acts as florigen, promotes the phase transition of axillary meristems, whereas BRANCHED1 (BRC1) antagonizes the florigen action in axillary buds. Here, we present evidences for another possible role of florigen in axillary bud development. Ectopic overexpression of FT or another florigen gene TWIN SISTER OF FT (TSF) with LEAFY (LFY) induces ectopic buds at cotyledonary axils, confirming the previous proposal that these genes are involved in formation of axillary buds. Taken together with our previous report that florigen promotes axillary shoot elongation, we propose that florigen regulates axillary bud development at multiple stages to coordinate it with flowering in Arabidopsis.     

A map-based cloning strategy employing a residual heterozygous line reveals that the GIGANTEA gene is involved in soybean maturity and flowering

Flowering is indicative of the transition from vegetative to reproductive phase, a critical event in the life cycle of plants. In soybean (Glycine max), a flowering quantitative trait locus, FT2, corresponding to the maturity locus E2, was detected in recombinant inbred lines (RILs) derived from the varieties "Misuzudaizu" (ft2/ft2; JP28856) and "Moshidou Gong 503" (FT2/FT2; JP27603). A map-based cloning strategy using the progeny of a residual heterozygous line (RHL) from the RIL was employed to isolate the gene responsible for this quantitative trait locus. A GIGANTEA ortholog, GmGIa (Glyma10g36600), was identified as a candidate gene. A common premature stop codon at the 10th exon was present in the Misuzudaizu allele and in other near isogenic lines (NILs) originating from Harosoy (e2/e2; PI548573). Furthermore, a mutant line harboring another premature stop codon showed an earlier flowering phenotype than the original variety, Bay (E2/E2; PI553043). The e2/e2 genotype exhibited elevated expression of GmFT2a, one of the florigen genes that leads to early flowering. The effects of the E2 allele on flowering time were similar among NILs and constant under high (43°N) and middle (36°N) latitudinal regions in Japan. These results indicate that GmGIa is the gene responsible for the E2 locus and that a null mutation in GmGIa may contribute to the geographic adaptation of soybean.     

Developing a method for customized induction of flowering

Background  

The ability to induce flowering on demand is of significant biotechnological interest. FT protein has been recently identified as an important component of the mobile flowering hormone, florigen, whose function is conserved across the plant kingdom. We therefore focused on manipulation of both endogenous and heterologous FT genes to develop a floral induction system where flowering would be inhibited until it was induced on demand. The concept was tested in the model plant Arabidopsis thaliana (Arabidopsis).     

Comparison of peptides in the phloem sap of flowering and non-flowering Perilla and lupine plants using microbore HPLC followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Physiological evidence indicates that flower formation is hormonally controlled. The floral stimulus, or florigen, is formed in the leaves as a response to an inductive photoperiod and translocated through the phloem to the apical meristem. However, because of difficulties in obtaining and analyzing phloem sap and the lack of a bioassay, the chemical nature of this stimulus is one of the major unsolved problems in plant biology. A combination of microbore high-performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was used to compare the contents of the phloem sap from flowering and non-flowering plants. Instead of using one- or two-dimensional gel electrophoresis, microbore HPLC separations allowed us to detect proteins/peptides that were very small and present at very low levels. We detected more than 100 components in the phloem sap of Perilla ocymoides L. and Lupinus albusL. Sequences for 16 peptides in a mass range from 1 to 9 kDa were obtained. Two of these could be identified, 11 showed similarity to known or deduced protein sequences, and three showed no similarity to any known protein or translated gene sequence. Four of these peptides were specific to, modified, or increased in plants that were flowering, indicating their possible role in flower induction. The sequences of these peptides showed similarities to two purine permeases, a protein with similarity to protein kinases, and a protein with no similarities to any known protein.     

The Medicago FLOWERING LOCUS T homolog, MtFTa1, is a key regulator of flowering time

FLOWERING LOCUS T (FT) genes encode proteins that function as the mobile floral signal, florigen. In this study, we characterized five FT-like genes from the model legume, Medicago (Medicago truncatula). The different FT genes showed distinct patterns of expression and responses to environmental cues. Three of the FT genes (MtFTa1, MtFTb1, and MtFTc) were able to complement the Arabidopsis (Arabidopsis thaliana) ft-1 mutant, suggesting that they are capable of functioning as florigen. MtFTa1 is the only one of the FT genes that is up-regulated by both long days (LDs) and vernalization, conditions that promote Medicago flowering, and transgenic Medicago plants overexpressing the MtFTa1 gene flowered very rapidly. The key role MtFTa1 plays in regulating flowering was demonstrated by the identification of fta1 mutants that flowered significantly later in all conditions examined. fta1 mutants do not respond to vernalization but are still responsive to LDs, indicating that the induction of flowering by prolonged cold acts solely through MtFTa1, whereas photoperiodic induction of flowering involves other genes, possibly MtFTb1, which is only expressed in leaves under LD conditions and therefore might contribute to the photoperiodic regulation of flowering. The role of the MtFTc gene is unclear, as the ftc mutants did not have any obvious flowering-time or other phenotypes. Overall, this work reveals the diversity of the regulation and function of the Medicago FT family.     

Historical perspective on breakthroughs in flowering field

Plants have evolved a mechanism to synchronize flowering time in response to environments. How plants recognize specific seasons for flowering has been a long sought question, thus, more than 100 years of research has been focused on this question. Especially in the past two decades, remarkable achievements have been made in identifying the molecular mechanism for flowering. Here we summarize the breakthroughs made in this field over the past century including discoveries of photoperiodic and vernalization-induced flowering, the identification of complex genetic pathways, and the recently proposed identity of florigen. In addition, we present the currently accepted model for a molecular mechanism toward flowering.