A MADS box gene, FLF (for FLOWERING LOCUS F ), isolated from a late-flowering, T-DNA-tagged Arabidopsis mutant, is a semidominant gene encoding a repressor of flowering. The FLF gene appears to integrate the vernalization-dependent and autonomous flowering pathways because its expression is regulated by genes in both pathways. The level of FLF mRNA is downregulated by vernalization and by a decrease in genomic DNA methylation, which is consistent with our previous suggestion that vernalization acts to induce flowering through changes in gene activity that are mediated through a reduction in DNA methylation. The flf-1 mutant requires a greater than normal amount of an exogenous gibberellin (GA3) to decrease flowering time compared with the wild type or with vernalization-responsive late-flowering mutants, suggesting that the FLF gene product may block the promotion of flowering by GAs. FLF maps to a region on chromosome 5 near the FLOWERING LOCUS C gene, which is a semidominant repressor of flowering in late-flowering ecotypes of Arabidopsis.

R ecently, we reported on a gene, FLOWERING LOCUS F (FLF), from Arabidopsis whose activity, as determined by steady-state mRNA levels, is correlated quantitatively with the period taken to initiate flowering (1). The FLF gene encodes a putative transcription factor of the MADS-box class, which, we concluded, acts as a repressor of floral induction. Genetic analyses of Arabidopsis ecotypes previously identified FLOW-ERING LOCUS C (FLC) as a semidominant repressor of floral induction (2-4). Because FLF has similar properties to FLC and maps to the same chromosomal region, the possibility arose that FLF and FLC are one and the same gene; this hypothesis has been confirmed by Michaels and Amasino (5). We suggest that the gene be referred to as FLC and the mutants flf-1 to flf-9 now be referred to as flc-11 to flc-19.In many plant species, flowering is promoted by a period of exposure to low temperature through a process known as vernalization. A number of the late-flowering mutants and ecotypes of Arabidopsis have a vernalization response. We found that the vernalization-responsive late-flowering mutants had a higher level of FLC transcript than was present in the wild types and, importantly, that the non-vernalization-responsive lateflowering mutants did not have increased FLC transcript levels. The vernalization-responsive ecotype C24, which is somewhat late-flowering, also has a high level of FLC transcript that is reduced by the low-temperature treatment (1). These observations suggested to us that FLC must be directly involved in the vernalization response.We have shown that reduction in the level of genomic methylation largely substitutes for the low-temperature treatment in promoting flowering (6-8), implicating a reduction in DNA methylation level as a component in the vernalization response. Early-flowering METHYLTRANSFERASE1 antisense plants, with a decreased level of genomic methylation, have a reduced level of FLC activity as compared with the nontransformed control (1). The correlation between FLC transcription and methylation levels implies that FLC is regulated by methylation status, but we are not able to say whether this is through an alteration of the methylation status of the FLC gene itself, or whether it involves other genes that regulate FLC expression.In this paper, we demonstrate that the FLC gene is involved directly in controlling the response to vernalization and more generally in the timing of the transition to the reproductive phase of development. We find that a reduction in FLC activity is a key component of the vernalization response in six late-flowering vernalization-responsive mutants and in the C24 and Pitztal ecotypes. The late-flowering mutants are mutated in genes that regulate the activity of the FLC gene. We further show that the quantitative responses in the promotion of flowering to various periods of exposure to low temperature are paralleled by the levels of FLC transcript, and that the changes observed at the transcriptional level are reflected in the level of the FLC pr...

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R ecently, we reported on a gene, FLOWERING LOCUS F (FLF), from Arabidopsis whose activity, as determined by steady-state mRNA levels, is correlated quantitatively with the period taken to initiate flowering (1). The FLF gene encodes a putative transcription factor of the MADS-box class, which, we concluded, acts as a repressor of floral induction. Genetic analyses of Arabidopsis ecotypes previously identified FLOW-ERING LOCUS C (FLC) as a semidominant repressor of floral induction (2-4). Because FLF has similar properties to FLC and maps to the same chromosomal region, the possibility arose that FLF and FLC are one and the same gene; this hypothesis has been confirmed by Michaels and Amasino (5). We suggest that the gene be referred to as FLC and the mutants flf-1 to flf-9 now be referred to as flc-11 to flc-19.In many plant species, flowering is promoted by a period of exposure to low temperature through a process known as vernalization. A number of the late-flowering mutants and ecotypes of Arabidopsis have a vernalization response. We found that the vernalization-responsive late-flowering mutants had a higher level of FLC transcript than was present in the wild types and, importantly, that the non-vernalization-responsive lateflowering mutants did not have increased FLC transcript levels. The vernalization-responsive ecotype C24, which is somewhat late-flowering, also has a high level of FLC transcript that is reduced by the low-temperature treatment (1). These observations suggested to us that FLC must be directly involved in the vernalization response.We have shown that reduction in the level of genomic methylation largely substitutes for the low-temperature treatment in promoting flowering (6-8), implicating a reduction in DNA methylation level as a component in the vernalization response. Early-flowering METHYLTRANSFERASE1 antisense plants, with a decreased level of genomic methylation, have a reduced level of FLC activity as compared with the nontransformed control (1). The correlation between FLC transcription and methylation levels implies that FLC is regulated by methylation status, but we are not able to say whether this is through an alteration of the methylation status of the FLC gene itself, or whether it involves other genes that regulate FLC expression.In this paper, we demonstrate that the FLC gene is involved directly in controlling the response to vernalization and more generally in the timing of the transition to the reproductive phase of development. We find that a reduction in FLC activity is a key component of the vernalization response in six late-flowering vernalization-responsive mutants and in the C24 and Pitztal ecotypes. The late-flowering mutants are mutated in genes that regulate the activity of the FLC gene. We further show that the quantitative responses in the promotion of flowering to various periods of exposure to low temperature are paralleled by the levels of FLC transcript, and that the changes observed at the transcriptional level are reflected in the level of the FLC pr...

Vernalization, the promotion of flowering by a prolonged period of low temperature, results in repression of the floral repressor FLOWERING LOCUS C ( FLC ) and in early flowering. This repression bears the hallmark of an epigenetic event: the low expression state is maintained over many cell division cycles, but expression is derepressed in progeny. We show that the two stages of the response of FLC to vernalization, the repression of FLC and the maintenance of the repression during growth at normal temperatures after vernalization, are mediated through different regions of the FLC gene. Both promoter and intragenic regions are required for the responses. We also identify a 75-bp region in the FLC promoter that, in addition to intragenic sequences, is required for expression in nonvernalized plants.

Analysis of the functions of Short Vegetative Phase (SVP)-like MADS-box genes in barley (Hordeum vulgare) indicated a role in determining meristem identity. Three SVP-like genes are expressed in vegetative tissues of barley: Barley MADS1 (BM1), BM10, and Vegetative to Reproductive Transition gene 2. These genes are induced by cold but are repressed during floral development. Ectopic expression of BM1 inhibited spike development and caused floral reversion in barley, with florets at the base of the spike replaced by tillers. Head emergence was delayed in plants that ectopically express BM1, primarily by delayed development after the floral transition, but expression levels of the barley VRN1 gene (HvVRN1) were not affected. Ectopic expression of BM10 inhibited spike development and caused partial floral reversion, where florets at the base of the spike were replaced by inflorescence-like structures, but did not affect heading date. Floral reversion occurred more frequently when BM1 and BM10 ectopic expression lines were grown in short-day conditions. BM1 and BM10 also inhibited floral development and caused floral reversion when expressed in Arabidopsis (Arabidopsis thaliana). We conclude that SVP-like genes function to suppress floral meristem identity in winter cereals.During the life cycle of a plant, the shoot apical meristem progresses through three phases of development: vegetative, inflorescence, and floral (Poethig, 1990). In each phase, the apical meristem produces a different set of organs. The vegetative meristem produces leaves, the inflorescence meristem produces leaves and floral meristems to form the inflorescence, and the floral meristem produces the organs that form the flower. The different phases of meristem development are controlled by genes that establish and maintain meristem identity.The shift from vegetative to inflorescence meristem identity, the floral transition, marks the beginning of the reproductive growth phase and is an important determinant of flowering time. In Arabidopsis (Arabidopsis thaliana), the Short Vegetative Phase (SVP) gene encodes a MADS-box transcription factor that delays the floral transition (Hartmann et al., 2000). Mutations that disrupt SVP cause early flowering (Hartmann et al., 2000), whereas ectopic expression of SVP results in late flowering. Ectopic expression of SVP also inhibits floral meristem identity, causing floral abnormalities such as the conversion of sepals and petals to leaf-like structures (Brill and Watson, 2004;Masiero et al., 2004) and causing inflorescence-like structures to develop within flowers (Brill and Watson, 2004). The development of inflorescences within flowers indicates that meristematic cells within the flower have lost floral identity and have formed an inflorescence instead of floral organs, a phenomenon known as floral reversion (Tooke et al., 2005). Presumably, ectopic expression of SVP causes floral reversion by interfering with a mechanism that maintains floral meristem identity.The Arabidopsis gene, AGAMOUS-LIKE 24 (AGL24)...

We have identified three Arabidopsis genes with GAMYB-like activity, AtMYB33, AtMYB65, and AtMYB101, which can substitute for barley (Hordeum vulgare) GAMYB in transactivating the barley ␣-amylase promoter. We have investigated the relationships between gibberellins (GAs), these GAMYB-like genes, and petiole elongation and flowering of Arabidopsis. Within 1 to 2 d of transferring plants from short-to long-day photoperiods, growth rate and erectness of petioles increased, and there were morphological changes at the shoot apex associated with the transition to flowering. These responses were accompanied by accumulation of GAs in the petioles (GA 1 by 11-fold and GA 4 by 3-fold), and an increase in expression of AtMYB33 at the shoot apex. Inhibition of GA biosynthesis using paclobutrazol blocked the petiole elongation induced by long days. Causality was suggested by the finding that, with GA treatment, plants flowered in short days, AtMYB33 expression increased at the shoot apex, and the petioles elongated and grew erect. That AtMYB33 may mediate a GA signaling role in flowering was supported by its ability to bind to a specific 8-bp sequence in the promoter of the floral meristem-identity gene, LEAFY, this same sequence being important in the GA response of the LEAFY promoter. One or more of these AtMYB genes may also play a role in the root tip during germination and, later, in stem tissue. These findings extend our earlier studies of GA signaling in the Gramineae to include a dicot species, Arabidopsis, and indicate that GAMYB-like genes may mediate GA signaling in growth and flowering responses.Gibberellins (GAs) regulate many aspects of plant growth and development. In the seed and seedling these include the production of hydrolytic enzymes, germination, and growth. In the adult plant, GAs are important in leaf and stem elongation, flowering, anther development, and fruit set (Pharis and King, 1985).Two classes of mutants have contributed much to an understanding of GA action (Thornton et al., 1999). One class includes dwarf mutants that are defective in GA biosynthesis. The other class includes response mutants such as Arabidopsis spindly (spy), GA-insensitive (gai), repressor of GA1-3 (rga), and the rice (Oryza sativa) d1 mutant. Many of the genes defined by these mutants have been cloned, but their molecular role in GA signaling is not yet fully understood (Jacobsen et al., 1996; Peng et al., 1997; Silverstone et al., 1998; Ashikari et al., 1999).An alternative approach to understanding GA signal transduction has involved functional studies, particularly with aleurone cells of cereals. These studies have identified a number of early GA signaling steps that precede expression of hydrolytic enzymes such as ␣-amylase. These steps involve heterotrimeric G-proteins (Jones et al., 1998; Ueguchi-Tanaka et al., 2000) and cGMP (Penson et al., 1996), which may in turn control the barley (Hordeum vulgare) HvGAMYB gene, whose expression is induced by GAs (Gubler et al., 1995).HvGAMYB encodes a transcriptional act...

Summary FLOWERING LOCUS C (FLC) inArabidopsis encodes a dosage dependent repressor of¯owering. We isolated ®ve FLC-related sequences from Brassica napus (BnFLC1±5). Expression of each of the ®ve sequences in Arabidopsis delayed¯owering signi®cantly, with the delay in¯owering time ranging from 3 weeks to more than 7 months, relative to the¯owering time of 3 weeks in untransformed Ler. In the reciprocal experiment, expression of Arabidopsis FLC (AtFLC) in an early¯owering B. napus cultivar delayed¯owering by 2±6 weeks, con®rming the requirement of this gene for¯oral repression. In B. napus, we show that late¯owering and responsiveness to vernalization correlate with the level of BnFLC mRNA expression. The different BnFLC genes show differential expression in leaves, stems and shoot tips, but expression is not detectable in roots. Vernalization dramatically reduces the level of BnFLC transcript and restores early¯owering in the winter cultivar Colombus. We conclude that BnFLC genes confer winter requirement in B. napus and account for the major vernalization-responsivē owering time differences in the different cultivars of B. napus in a manner analogous to that of AtFLC in Arabidopsis ecotypes.