Cadmium induces early flowering in Arabidopsis

We found that cadmium promoted flowering in Arabidopsis and suppressed nitric oxide accumulation in leaves. Supplementation with NO donor SNP delayed flowering, whereas application of NO scavenger cPTIO further promoted the transition from vegetative to reproductive stage under Cd stress. Semi-quantitative RT-PCR showed that Cd treatment up-regulated the expression of CONSTANS and FLOWERING LOCUS T, whereas down-regulated the expression of FLOWERING LOCUS C.     

Analysis of flowering pathway integrators in Arabidopsis

Flowering is regulated by an integrated network of several genetic pathways in Arabidopsis. The key genes integrating multiple flowering pathways are FT, SOC1 and LFY. To elucidate the interactions among these integrators, genetic analyses were performed. FT and SOC1 share the common upstream regulators CO, a key component in the long day pathway, and FLC, a flowering repressor integrating autonomous and vernalization pathways. However, the soc1 mutation further delayed the flowering time of long day pathway mutants including ft, demonstrating that SOC1 acts partially independently of FT. Although soc1 did not show an obvious defect in flower meristem determination on its own, it dramatically increased the number of coflorescences in a lfy mutant, which is indicative of a defect in floral initiation. Therefore, double mutant analysis shows that the three integrators have both overlapping and independent functions in the determination of flowering time and floral initiation. The expression analysis showed that FT regulates SOC1 expression, and SOC1 regulates LFY expression, but not vice versa, which is consistent with the fact that FT and LFY have the least overlapping functions among the three integrators. The triple mutation ft soc1 lfy did not block flowering completely under long days, indicating the presence of other integrators. Finally, vernalization accelerated flowering of flc ft soc1 and ft soc1 lfy triple mutants, which shows that the vernalization pathway also has targets other than FLC, FT, SOC1 and LFY. Our genetic analysis reveals the intricate nature of genetic networks for flowering.     

Integration of flowering signals in winter-annual Arabidopsis

Photoperiod is the primary environmental factor affecting flowering time in rapid-cycling accessions of Arabidopsis (Arabidopsis thaliana). Winter-annual Arabidopsis, in contrast, have both a photoperiod and a vernalization requirement for rapid flowering. In winter annuals, high levels of the floral inhibitor FLC (FLOWERING LOCUS C) suppress flowering prior to vernalization. FLC acts to delay flowering, in part, by suppressing expression of the floral promoter SOC1 (SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1). Vernalization leads to a permanent epigenetic suppression of FLC. To investigate how winter-annual accessions integrate signals from the photoperiod and vernalization pathways, we have examined activation-tagged alleles of FT and the FT homolog, TSF (TWIN SISTER OF FT), in a winter-annual background. Activation of FT or TSF strongly suppresses the FLC-mediated late-flowering phenotype of winter annuals; however, FT and TSF overexpression does not affect FLC mRNA levels. Rather, FT and TSF bypass the block to flowering created by FLC by activating SOC1 expression. We have also found that FLC acts as a dosage-dependent inhibitor of FT expression. Thus, the integration of flowering signals from the photoperiod and vernalization pathways occurs, at least in part, through the regulation of FT, TSF, and SOC1.     

Revisiting phase transition during flowering in Arabidopsis

Single-phase transition during flowering has been suggested by Hempel and Feldman (1994) [Planta 192: 276]. When early flowering ecotypes of Arabidopsis were microscopically observed, a long day signal simultaneously induced the acropetal (bottom to top) production of flower primordia and the basipetal (top to bottom) differentiation of paraclades (axillary flowering shoots) from the axils of pre-existing leaf primordia. However, this model could not account for the production of an extra number of secondary shoots in the TERMINAL FLOWER 1 overexpressor line or AGL20 overexpressor line in Columbia background with a functional allele of FRIGIDA. We report here that Columbia with a functional allele of FRIGIDA under long days and Columbia under short days show an inflorescence-producing phase between the vegetative and the flower-producing phases, supporting two-step phase transition during flowering. In addition, a late-flowering mutant, fwa shows an inflorescence phase but fca, fy and fve follow a single-phase transition, suggesting flowering time mutations have different effects on phase transition during flowering.     

Association of glutathione with flowering in Arabidopsis thaliana

In order to study the relationship between GSH and flowering, wild-type and late-flowering mutant, fca-1, of Arabidopsis thaliana were treated with L-buthionine sulfoximine (BSO), a specific inhibitor of GSH biosynthesis, under long-day conditions. BSO treatment of the fca-1 mutant starting at 17 d after imbibition promoted flowering. However, when the treatment was started at 12 d after imbibition, BSO treatment at 10(-4) M resulted in an inhibition of flowering. This inhibitory effect of BSO on flowering was abolished by GSH treatment at 10(-4) M, although GSH treatment at an increased concentration of 10(-3) M clearly delayed flowering. In contrast, BSO treatment of wild-type plants starting at 12 d after imbibition promoted flowering, whose effect was abolished by GSH application. In the fca-1 mutant, whose endogenous GSH levels were high, chilling treatment lowered the GSH levels and promoted flowering, as was the case in the BSO treatment. An A. thaliana mutant, cad2-1, which has a defect in GSH biosynthesis also exhibited late flowering. The late-flowering phenotype of this mutant tended to be strengthened by BSO and abolished by GSH treatment. These results suggest that flowering is associated with the rate of GSH biosynthesis and/or the levels of GSH in A. thaliana.     

Specification of chimeric flowering shoots in wild-type Arabidopsis

Within wild-type Arabidopsis populations, a subset of the plants were found to have a single chimeric shoot on their primary shoot axes. The chimeric shoots were located below the lowest primary-axis flower; and they exhibited features of both flowers and paraclades (lateral flowering shoots). Morphological analyses of chimeric shoots indicated that they developed from single primordia. In each chimeric shoot, the side furthest from the apical meristem was specified as 'flower'—while the side closest to the meristem was specified as 'paraclade'—suggesting that a stimulus from outside the apical meristem can directly induce primordia to develop as flowers. It is concluded that the development of the teratological chimeric shoots resulted from the overlap of the vegetative and floral specification processes within single primordia.     

Conservation of Arabidopsis flowering genes in model legumes

The model plants Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa) have provided a wealth of information about genes and genetic pathways controlling the flowering process, but little is known about the corresponding pathways in legumes. The garden pea (Pisum sativum) has been used for several decades as a model system for physiological genetics of flowering, but the lack of molecular information about pea flowering genes has prevented direct comparison with other systems. To address this problem, we have searched expressed sequence tag and genome sequence databases to identify flowering-gene-related sequences from Medicago truncatula, soybean (Glycine max), and Lotus japonicus, and isolated corresponding sequences from pea by degenerate-primer polymerase chain reaction and library screening. We found that the majority of Arabidopsis flowering genes are represented in pea and in legume sequence databases, although several gene families, including the MADS-box, CONSTANS, and FLOWERING LOCUS T/TERMINAL FLOWER1 families, appear to have undergone differential expansion, and several important Arabidopsis genes, including FRIGIDA and members of the FLOWERING LOCUS C clade, are conspicuously absent. In several cases, pea and Medicago orthologs are shown to map to conserved map positions, emphasizing the closely syntenic relationship between these two species. These results demonstrate the potential benefit of parallel model systems for an understanding of flowering phenology in crop and model legume species.     

Interaction of Polycomb-group proteins controlling flowering in Arabidopsis

In Arabidopsis, the EMBYRONIC FLOWER2 (EMF2), VERNALISATION2 (VRN2) and FERTILISATION INDEPENDENT ENDOSPERM2 (FIS2) genes encode related Polycomb-group (Pc-G) proteins. Their homologues in animals act together with other Pc-G proteins as part of a multimeric complex, Polycomb Repressive Complex 2 (PRC2), which functions as a histone methyltransferase. Despite similarities between the fis2 mutant phenotype and those of some other plant Pc-G members, it has remained unclear how the FIS2/EMF2/VRN2 class Pc-G genes interact with the others. We have identified a weak emf2 allele that reveals a novel phenotype with striking similarity to that of severe mutations in another Pc-G gene, CURLY LEAF (CLF), suggesting that the two genes may act in a common pathway. Consistent with this, we demonstrate that EMF2 and CLF interact genetically and that this reflects interaction of their protein products through two conserved motifs, the VEFS domain and the C5 domain. We show that the full function of CLF is masked by partial redundancy with a closely related gene, SWINGER (SWN), so that null clf mutants have a much less severe phenotype than emf2 mutants. Analysis in yeast further indicates a potential for the CLF and SWN proteins to interact with the other VEFS domain proteins VRN2 and FIS2. The functions of individual Pc-G members may therefore be broader than single mutant phenotypes reveal. We suggest that plants have Pc-G protein complexes similar to the Polycomb Repressive Complex2 (PRC2) of animals, but the duplication and subsequent diversification of components has given rise to different complexes with partially discrete functions.     

The role of cryptochrome 2 in flowering in Arabidopsis

We have investigated the genetic interactions between cry2 and the various flowering pathways in relation to the regulation of flowering by photoperiod and vernalization. For this, we combined three alleles of CRY2, the wild-type CRY2-Landsberg erecta (Ler), a cry2 loss-of-function null allele, and the gain-of-function CRY2-Cape Verde Islands (Cvi), with mutants representing the various photoreceptors and flowering pathways. The analysis of CRY2 alleles combined with photoreceptor mutants showed that CRY2-Cvi could compensate the loss of phyA and cry1, also indicating that cry2 does not require functional phyA or cry1. The analysis of mutants of the photoperiod pathway showed epistasis of co and gi to the CRY2 alleles, indicating that cry2 needs the product of CO and GI genes to promote flowering. All double mutants of this pathway showed a photoperiod response very much reduced compared with Ler. In contrast, mutations in the autonomous pathway genes were additive to the CRY2 alleles, partially overcoming the effects of CRY2-Cvi and restoring day length responsiveness. The three CRY2 alleles were day length sensitive when combined with FRI-Sf2 and/or FLC-Sf2 genes, which could be reverted when the delay of flowering caused by FRI-Sf2 and FLC-Sf2 alleles was removed by vernalization. In addition, we looked at the expression of FLC and CRY2 genes and showed that CRY2 is negatively regulated by FLC. These results indicate an interaction between the photoperiod and the FLC-dependent pathways upstream to the common downstream targets of both pathways, SOC1 and FT.     

Extensive phenotypic variation in early flowering mutants of Arabidopsis

Flowering time, the major regulatory transition of plant sequential development, is modulated by multiple endogenous and environmental factors. By phenotypic profiling of 80 early flowering mutants of Arabidopsis, we examine how mutational reduction of floral repression is associated with changes in phenotypic plasticity and stability. Flowering time measurements in mutants reveal deviations from the linear relationship between the number of leaves and number of days to bolting described for natural accessions and late flowering mutants. The deviations correspond to relative early bolting and relative late bolting phenotypes. Only a minority of mutants presents no detectable phenotypic variation. Mutants are characterized by a broad release of morphological pleiotropy under short days, with leaf characters being most variable. They also exhibit changes in phenotypic plasticity across environments for florigenic-related responses, including the reaction to light and dark, photoperiodic behavior, and Suc sensitivity. Morphological pleiotropy and plasticity modifications are differentially distributed among mutants, resulting in a large diversity of multiple phenotypic changes. The pleiotropic effects observed may indicate that floral repression defects are linked to global developmental perturbations. This first, to our knowledge, extensive characterization of phenotypic variation in early flowering mutants correlates with the reports that most factors recruited in floral repression at the molecular genetic level correspond to ubiquitous regulators. We discuss the importance of functional ubiquity for floral repression with respect to robustness and flexibility of network biological systems.     

Diversity of flowering responses in wild Arabidopsis thaliana strains

Although multiple environmental cues regulate the transition to flowering in Arabidopsis thaliana, previous studies have suggested that wild A. thaliana accessions fall primarily into two classes, distinguished by their requirement for vernalization (extended winter-like temperatures), which enables rapid flowering under long days. Much of the difference in vernalization response is apparently due to variation at two epistatically acting loci, FRI and FLC. We present the response of over 150 wild accessions to three different environmental variables. In long days, FLC is among those genes whose expression is most highly correlated with flowering. In short days, FRI and FLC are less important, although their contribution is still significant. In addition, there is considerable variation not only in vernalization response, but also in the response to differences in day length or ambient growth temperature. The identification of accessions that flower relatively early or late in specific environments suggests that many of the flowering-time pathways identified by mutagenesis, such as those that respond to day length, contribute to flowering-time variation in the wild. In contrast to differences in vernalization requirement, which are mainly mediated by FRI and FLC, it seems that variation in these other pathways is due to allelic effects at several different loci.     

Mutations in AP22.65 accelerate flowering in Arabidopsis thaliana

Identification of the gene(s) responsible for flowering time in Arabidopsis has significant implications. We used the T-DNA insertion library of Arabidopsis thaliana to screen an early-flowering mutant that exhibits accelerated flowering under short-day conditions. AP22.65, a novel flowering-time gene in that species, was isolated and identified via genome-walking and bioinformatics analysis. The flowering time of AP22.65-complementing plants was similar to that of the Col-0 wild type (WT). Conversely, its overexpression delayed flowering. Consistent with this phenotype, expression of AP22.65 was decreased in the ap22.65-1 mutant, recovered in AP22.65-complementing plants, and increased in AP22.65-overexpressing plants. Compared with the WT, expression levels of critical genes in different flowering pathways, i.e., SPY, FLC, GI, CO, FT, and LFY, were down-regulated in loss-of-function mutants. Expression of AP22.65 was distributed in flowers, siliques, rosette leaves, and whole seedlings. Therefore, this gene may be a negative regulator of Arabidopsis flowering.     

CRYPTOCHROME2 in vascular bundles regulates flowering in Arabidopsis

Plants make full use of light signals to determine the timing of flowering. In Arabidopsis thaliana, a blue/UV-A photoreceptor, CRYPTOCHROME 2 (cry2), and a red/far-red photoreceptor, PHYTOCHROME B (phyB), are two major photoreceptors that control flowering. The light stimuli for the regulation of flowering are perceived by leaves. We have recently shown that phyB expression in mesophyll but not in vascular bundles suppresses the expression of a key flowering regulator, FLOWERING LOCUS T (FT), in vascular bundles. In this study, we asked where in the leaf cry2 perceives light stimuli to regulate flowering. To answer this question, we established transgenic Arabidopsis lines in which the cry2-green fluorescent protein (GFP) fusion was expressed under the control of organ/tissue-specific promoters in a cry2-deficient mutant background. Analysis of these lines revealed that expression of cry2-GFP in vascular bundles, but not in epidermis or mesophyll, rescued the late flowering phenotype. We further confirmed that cry2-GFP expressed in vascular bundles increased FT expression only in vascular bundles. Hence, in striking contrast with phyB, cry2 most likely regulates FT expression in a cell-autonomous manner.     

Regulation of flowering time by Arabidopsis MSI1

The transition to flowering is tightly controlled by endogenous programs and environmental signals. We found that MSI1 is a novel flowering-time gene in Arabidopsis. Both partially complemented msi1 mutants and MSI1 antisense plants were late flowering, whereas ectopic expression of MSI1 accelerated flowering. Physiological experiments revealed that MSI1 is similar to genes from the autonomous promotion of flowering pathway. Expression of most known flowering-time genes did not depend on MSI1, but the induction of SOC1 was delayed in partially complemented msi1 mutants. Delayed activation of SOC1 is often caused by increased expression of the floral repressor FLC. However, MSI1 function is independent of FLC. MSI1 is needed to establish epigenetic H3K4 di-methylation and H3K9 acetylation marks in SOC1 chromatin. The presence of these modifications correlates with the high levels of SOC1 expression that induce flowering in Arabidopsis. Together, the control of flowering time depends on epigenetic mechanisms for the correct expression of not only the floral repressor FLC, but also the floral activator SOC1.     

FLC: A key regulator of flowering time in Arabidopsis

The timing of floral transition has significant consequences for reproductive success in plants. The molecular genetic dissection of flowering time control in Arabidopsis identified an integrated network of pathways that quantitatively control this developmental switch. A central player in this process is the FLOWERING LOCUS C gene (FLC), which blocks flowering by inhibiting the genes required to switch the meristem from vegetative to floral development. Three systems (the FRIGIDA gene, vernalization, and the autonomous pathway) all influence the state of FLC. Last years many new genes have been identified that regulate FLC expression, and most of them are involved in the modification of FLC chromatin. This review focuses on recent insights in FLC regulation.