The Function of Florigen in the Vegetative-to-Reproductive Phase Transition in and around the Shoot Apical Meristem.

Plants undergo a series of developmental phases throughout their life-cycle, each characterized by specific processes. Three critical features distinguish these phases: the arrangement of primordia (phyllotaxis), the timing of their differentiation (plastochron) and the characteristics of the lateral organs and axillary meristems. Identifying the unique molecular features of each phase, determining the molecular triggers that cause transitions and understanding the molecular mechanisms underlying these transitions are keys to gleaning a complete understanding of plant development. During the vegetative phase, the shoot apical meristem (SAM) facilitates continuous leaf and stem formation, with leaf development as the hallmark. The transition to the reproductive phase induces significant changes in these processes, driven mainly by the protein FT (FLOWERING LOCUS T) in Arabidopsis and proteins encoded by FT orthologs, which are specified as 'florigen'. These proteins are synthesized in leaves and transported to the SAM, and act as the primary flowering signal, although its impact varies among species. Within the SAM, florigen integrates with other signals, culminating in developmental changes. This review explores the central question of how florigen induces developmental phase transition in the SAM. Future research may combine phase transition studies, potentially revealing the florigen-induced developmental phase transition in the SAM.

PHOSPHORYLETHANOLAMINE CYTIDYLYLTRANSFERASE 1 modulates flowering in a florigen-independent manner by regulating SVP.

PHOSPHORYLETHANOLAMINE CYTIDYLYLTRANSFERASE 1 (PECT1) regulates phosphatidylethanolamine biosynthesis and controls the phosphatidylethanolamine:phosphatidylcholine ratio in Arabidopsis thaliana Previous studies have suggested that PECT1 regulates flowering time by modulating the interaction between phosphatidylcholine and FLOWERING LOCUS T (FT), a florigen, in the shoot apical meristem (SAM). Here, we show that knockdown of PECT1 by artificial microRNA in the SAM (pFD::amiR-PECT1) accelerated flowering under inductive and even non-inductive conditions, in which FT transcription is almost absent, and in ft-10 twin sister of ft-1 double mutants under both conditions. Transcriptome analyses suggested that PECT1 affects flowering by regulating SHORT VEGETATIVE PHASE (SVP) and GIBBERELLIN 20 OXIDASE 2 (GA20ox2). SVP misexpression in the SAM suppressed the early flowering of pFD::amiR-PECT1 plants. pFD::amiR-PECT1 plants showed increased gibberellin (GA) levels in the SAM, concomitant with the reduction of REPRESSOR OF GA1-3 levels. Consistent with this, GA treatment had little effect on flowering time of pFD::amiR-PECT1 plants and the GA antagonist paclobutrazol strongly affected flowering in these plants. Together, these results suggest that PECT1 also regulates flowering time through a florigen-independent pathway, modulating SVP expression and thus regulating GA production.

Genetics of phenological development and implications for seed yield in lentil.

Understanding phenology, its genetics and agronomic consequences, is critical for crop adaptation. Here we aim at (1) characterising lentil response to photoperiod with a focus on five loci: the lentil ELF3 ortholog Sn, two loci linked to clusters of lentil FT orthologs and two loci without candidates in chromosomes 2 and 5 (exp. 1: 36 lines, short and long day in phytotron); (2) establishing phenology-yield relationship (exp. 2: 25 lines, 11 field environments). A vintage perspective, where we quantify time trends in phenotype over three decades of breeding, links both experiments. Yield increased linearly from older to newer varieties at 29 kg ha-1 yr-1 or 1.5% yr-1, correlated negatively with flowering time in both winter- and summer-rainfall regimes, and decoupled from biomass in favourable environments. Time to flowering shortened from older to newer varieties at -0.56 % yr-1 in the field, and -0.42 % yr-1 (short day) and -0.99 % yr-1 (long day) in the phytotron. Early-flowering lines of diverse origin carried multiple early alleles for the five loci, indicating that at least some of these loci affect phenology additively. Current germplasm primarily features the early flowering haplotype for an FTb cluster region, hence the potential to increase phenological diversity with yield implications.

Multifunctional chemical inhibitors of the florigen activation complex discovered by structure-based high-throughput screening.

Structure-based high-throughput screening of chemical compounds that target protein-protein interactions (PPIs) is a promising technology for gaining insight into how plant development is regulated, leading to many potential agricultural applications. At present, there are no examples of using high-throughput screening to identify chemicals that target plant transcriptional complexes, some of which are responsible for regulating multiple physiological functions. Florigen, a protein encoded by FLOWERING LOCUS T (FT), was initially identified as a molecule that promotes flowering and has since been shown to regulate flowering and other developmental phenomena such as tuber formation in potato (Solanum tuberosum). FT functions as a component of the florigen activation complex (FAC) with a 14-3-3 scaffold protein and FD, a bZIP transcription factor that activates downstream gene expression. Although 14-3-3 is an important component of FAC, little is known about the function of the 14-3-3 protein itself. Here, we report the results of a high-throughput in vitro fluorescence resonance energy transfer (FRET) screening of chemical libraries that enabled us to identify small molecules capable of inhibiting FAC formation. These molecules abrogate the in vitro interaction between the 14-3-3 protein and the OsFD1 peptide, a rice (Oryza sativa) FD, by directly binding to the 14-3-3 protein. Treatment with S4, a specific hit molecule, strongly inhibited FAC activity and flowering in duckweed, tuber formation in potato, and branching in rice in a dose-dependent manner. Our results demonstrate that the high-throughput screening approach based on the three-dimensional structure of PPIs is suitable in plants. In this study, we have proposed good candidate compounds for future modification to obtain inhibitors of florigen-dependent processes through inhibition of FAC formation.

Cytokinin increases vegetative growth period by suppressing florigen expression in rice and maize.

Increasing the vegetative growth period of crops can increase biomass and grain yield. In rice (Oryza sativa), the concentration of trans -zeatin, an active cytokinin, was high in the leaves during vegetative growth and decreased rapidly upon induction of florigen expression, suggesting that this hormone is involved in the regulation of the vegetative phase. To elucidate whether exogenous cytokinin application influences the length of the vegetative phase, we applied 6-benzylaminopurine (BAP) to rice plants at various developmental stages. Our treatment delayed flowering time by 8-9 days when compared with mock-treated rice plants, but only at the transition stage when the flowering signals were produced. Our observations also showed that flowering in the paddy field is delayed by thidiazuron, a stable chemical that mimics the effects of cytokinin. The transcript levels of florigen genes Heading date 3a (Hd3a) and Rice Flowering locus T1 (RFT1) were significantly reduced by the treatment, but the expression of Early heading date 1 (Ehd1), a gene found directly upstream of the florigen genes, was not altered. In maize (Zea mays), similarly, BAP treatment increased the vegetative phage by inhibiting the expression of ZCN8, an ortholog of Hd3a. We showed that cytokinin treatment induced the expression of two type-A response regulators (OsRR1 and OsRR2) which interacted with Ehd1, a type-B response regulator. We also observed that cytokinin did not affect flowering time in ehd1 knockout mutants. Our study indicates that cytokinin application increases the duration of the vegetative phase by delaying the expression of florigen genes in rice and maize by inhibiting Ehd1.

Identification, characterization, and comprehensive expression profiling of floral master regulators in pigeon pea (Cajanus cajan [L.] Millspaugh).

Pigeon pea is an important protein-rich pulse crop. Identification of flowering master regulators in pigeon pea is highly imperative as indeterminacy and late flowering are impediments towards yield improvement. A genome-wide analysis was performed to explore flowering orthologous groups in pigeon pea. Among the 412 floral orthologs identified in pigeon pea, 148 genes belong to the meristem identity, photoperiod-responsive, and circadian clock-associated ortholog groups. Our comparative genomics study revealed purifying selection pressures (ka/ks) on floral orthologs, and duplication patterns and evolution through synteny with other model species. Phylogenetic analysis of floral genes substantiated a connection between pigeon pea plant architecture and flowering time as all the PEBP domain-containing genes belong to meristem identity floral networks of pigeon pea. Expression profiling of eleven major orthologs in contrasting determinate and indeterminate genotypes indicated that these orthologs might be involved in flowering regulation. Expression of floral inducer, FT, and floral repressor, TFL1, was non-comparable in indeterminate genotypes across all the developmental stages of pigeon pea. However, dynamic FT/TFL1 expression ratio detected in all tissues of both the genotypes suggested their role in floral transition. One TFL1 ortholog having high sequence conserveness across pigeon pea genotypes showed differential expression indicating genotype-dependent regulation of this ortholog. Presence of conserved 6mA-methylation patterns in light-responsive elements and in other cis-regulatory elements of FT and TFL1 across different plant genotypes indicated possible involvement of epigenetic regulation in flowering.

OsFTL12, a member of FT-like family, modulates the heading date and plant architecture by florigen repression complex in rice.

FLOWERING LOCUS T (FT), a florigen in Arabidopsis, plays critical roles in floral transition. Among 13 FT-like members in rice, OsFTL2 (Hd3a) and OsFTL3 (RFT1), two rice homologues of FT, have been well characterized to act as florigens to induce flowering under short-day (SD) and long-day (LD) conditions, respectively, but the functions of other rice FT-like members remain largely unclear. Here, we show that OsFTL12 plays an antagonistic function against Hd3a and RFT1 to modulate the heading date and plant architecture in rice. Unlike Hd3a and RFT1, OsFTL12 is not regulated by daylength and highly expressed in both SD and LD conditions, and delays the heading date under either SD or LD conditions. We further demonstrate that OsFTL12 interacts with GF14b and OsFD1, two key components of the florigen activation complex (FAC), to form the florigen repression complex (FRC) by competing with Hd3a for binding GF14b. Notably, OsFTL12-FRC can bind to the promoters of the floral identity genes OsMADS14 and OsMADS15 and suppress their expression. The osmads14 osmads15 double mutants could not develop panicles and showed erect leaves. Taken together, our results reveal that different FT-like members can fine-tune heading date and plant architecture by regulating the balance of FAC and FRC in rice.

The FLOWERING LOCUS T gene expression is controlled by high-irradiance response and external coincidence mechanism in long days in Arabidopsis.

In natural long days, the florigen gene FLOWERING LOCUS T (FT) shows a bimodal expression pattern with morning and dusk peaks in Arabidopsis. This pattern differs from the one observed in the laboratory, and little is known about underlying mechanisms. A red : far-red (R : FR) ratio difference between sunlight and fluorescent light causes this FT pattern mismatch. We showed that bimodal FT expression patterns were induced in a day longer than 14 h with sunlight R : FR (= c. 1) conditions. By circadian gating experiments, we found that cumulative exposure of R : FR-adjusted light (R : FR ratio was adjusted to 1 with FR supplement) spanning from the afternoon to the next morning required full induction of FT in the morning. Conversely, only 2 h of R : FR adjustment in the late afternoon was sufficient for FT induction at dusk. We identified that phytochrome A (phyA) is required for the morning FT expression in response to the R : FR adjustment on the previous day. As a part of this mechanism, we showed that PHYTOCHROME-INTERACTING FACTOR 7 contributes to FT regulation. Our results suggest that phyA-mediated high-irradiance response and the external coincidence mechanism contribute to morning FT induction under natural long-day conditions.

Homeobox transcription factors OsZHD1 and OsZHD2 induce inflorescence meristem activity at floral transition in rice.

Floral transition starts in the leaves when florigens respond to various environmental and developmental factors. Among several regulatory genes that are preferentially expressed in the inflorescence meristem during the floral transition, this study examines the homeobox genes OsZHD1 and OsZHD2 for their roles in regulating this transition. Although single mutations in these genes did not result in visible phenotype changes, double mutations in these genes delayed flowering. Florigen expression was not altered in the double mutants, indicating that the delay was due to a defect in florigen signaling. Morphological analysis of shoot apical meristem at the early developmental stage indicated that inflorescence meristem development was significantly delayed in the double mutants. Overexpression of ZHD2 causes early flowering because of downstream signals after the generation of florigens. Expression levels of the auxin biosynthesis genes were reduced in the mutants and the addition of indole-3-acetic acid recovered the defect in the mutants, suggesting that these homeobox genes play a role in auxin biosynthesis. A rice florigen, RICE FLOWERING LOCUS T 1, binds to the promoter regions of homeobox genes. These results indicate that florigens stimulate the expression of homeobox genes, enhancing inflorescence development in the shoot apex.

Quantitative Analysis of Florigen for the Variability of Floral Induction in Cabbage/Radish Inter-generic Grafting.

Grafting-induced flowering is a key phenomenon to understand systemic floral induction caused by florigen. It can also be used as a breeding technique enabling rapid seed production of crops with long generation times. However, the degree of floral induction in grafted plants is often variable. Moreover, it is difficult in some crop species. Here, we explored the factors promoting variability in the grafting-induced flowering of cabbage (Brassica oleracea L. var. capitata), an important vegetable crop with a long generation time, via the quantitative analysis of florigen accumulation. Significant variability in the flowering response of grafted cabbage was observed when rootstocks of different genotypes were used. As reported previously, B. oleracea rootstocks did not induce the flowering of grafted cabbage plants, but radish (Raphanus sativus L.) rootstocks unstably did, depending on the accessions used. Immunoblotting analysis of the FLOWERING LOCUS T (FT) protein, a main component of florigen, revealed that floral induction was quantitatively correlated with the level of accumulated FT protein in the grafted scion. To identify rootstock factors that cause variability in the floral induction of the grafted scion, we investigated FT protein accumulation and flowering response in grafted scions when the transcription levels of FT and the leaf area of rootstocks were altered by vernalization, daylength and leaf trimming treatments. We concluded that increasing the total amount of FT protein produced in the rootstock is important for the stable floral induction of the grafted cabbage, and this can be accomplished by increasing FT transcription and the leaf area of the rootstock.

Transient activity of the florigen complex during the floral transition in Arabidopsis thaliana.

FLOWERING LOCUS T (FT) is an essential component of florigen in Arabidopsis thaliana Transcription of FT is induced in leaves, and the resulting FT protein is transported to the shoot apex, in which it initiates floral development. Previous analyses suggest that, together with the b-ZIP transcription factor FD, FT regulates the transcription of downstream targets such as APETALA1 (AP1) in floral anlagen. However, conclusive in vivo evidence that FT is transported to the shoot apex to form an FT-FD complex is lacking. Here, using an innovative in vivo imaging technique, we show that the FT-FD complex and AP1 colocalise in floral anlagen. In addition, the FT-FD complex disappears soon after the floral transition owing to a reduction in FD transcripts in the shoot apex. We further show that misinduction of FD activity after the transition leads to defective reproductive development. Taken together, our results indicate that the FT-FD complex functions as a transient stimulus and imply that a regulatory mechanism exists during the floral transition that reduces FT-FD complex levels via modulation of FD expression.

Florigen and its homologs of FT/CETS/PEBP/RKIP/YbhB family may be the enzymes of small molecule metabolism: review of the evidence.

BACKGROUND: Flowering signals are sensed in plant leaves and transmitted to the shoot apical meristems, where the formation of flowers is initiated. Searches for a diffusible hormone-like signaling entity ("florigen") went on for many decades, until a product of plant gene FT was identified as the key component of florigen in the 1990s, based on the analysis of mutants, genetic complementation evidence, and protein and RNA localization studies. Sequence homologs of FT protein are found throughout prokaryotes and eukaryotes; some eukaryotic family members appear to bind phospholipids or interact with the components of the signal transduction cascades. Most FT homologs are known to share a constellation of five charged residues, three of which, i.e., two histidines and an aspartic acid, are located at the rim of a well-defined cavity on the protein surface. RESULTS: We studied molecular features of the FT homologs in prokaryotes and analyzed their genome context, to find tentative evidence connecting the bacterial FT homologs with small molecule metabolism, often involving substrates that contain sugar or ribonucleoside moieties. We argue that the unifying feature of this protein family, i.e., a set of charged residues conserved at the sequence and structural levels, is more likely to be an enzymatic active center than a catalytically inert ligand-binding site. CONCLUSIONS: We propose that most of FT-related proteins are enzymes operating on small diffusible molecules. Those metabolites may constitute an overlooked essential ingredient of the florigen signal.

The transcription factor OsGATA6 regulates rice heading date and grain number per panicle.

Heading date, panicle architecture, and grain size are key traits that affect the yield of rice (Oryza sativa). Here, we identified a new gene, OsGATA6, whose product regulates heading date. Overexpression of OsGATA6 resulted in delayed heading, increased grain number, and decreased grain size. Knockdown lines generated by artificial microRNA (amiRNA) and CRISPR genome-edited lines of OsGATA6 both showed earlier heading, decreased grain number, and increased grain size. These results suggested that OsGATA6 negatively regulates heading date, positively regulates panicle development, and affects grain size. OsGATA6 was found to be constitutively expressed in rice, and strongly expressed in young leaves and panicles. In situ hybridization analyses showed that OsGATA6 was specifically localized in superficial cells of the panicle primordium. Overexpression lines show decreased expression of RFT1 and Hd3a, which promote heading. OsMFT1, which delays heading date and increases grain number, was down-regulated in amiRNA lines. Further analyses showed that OsGATA6 could bind to the promoter of OsMFT1 and induce its expression, thereby regulating heading date and panicle development. Overexpression of OsGATA6 in Arabidopsis resulted in repressed expression of AtFT and late flowering, suggesting that its function is similar. Taken together, we have identified a new GATA regulator that influences rice heading date and grain number, which potentially increases rice yield.

The genetic architecture of flowering time changes in pea from wild to crop.

Change in phenology has been an important component in crop evolution, and selection for earlier flowering through a reduction in environmental sensitivity has helped broaden adaptation in many species. Natural variation for flowering in domesticated pea (Pisum sativum L.) has been noted and studied for decades, but there has been no clear account of change relative to its wild progenitor. Here we examined the genetic control of differences in flowering time between wild P. sativum ssp. humile and a typical late-flowering photoperiodic P. s. sativum accession in a recombinant inbred population under long and short photoperiods. Our results confirm the importance of the major photoperiod sensitivity locus Hr/PsELF3a and identify two other loci on chromosomes 1 (DTF1) and 3 (DTF3) that contribute to earlier flowering in the domesticated line under both photoperiods. The domesticated allele at a fourth locus on chromosome 6 (DTF6) delays flowering under long days only. Map positions, inheritance patterns, and expression analyses in near-isogenic comparisons imply that DTF1, DTF3, and DTF6 represent gain-of-function alleles of the florigen/antiflorigen genes FTa3, FTa1, and TFL1c/LF, respectively. This echoes similar variation in chickpea and lentil, and suggests a conserved route to reduced photoperiod sensitivity and early phenology in temperate pulses.

Genetic analysis of early phenology in lentil identifies distinct loci controlling component traits.

Modern-day domesticated lentil germplasm is generally considered to form three broad adaptation groups: Mediterranean, South Asian, and northern temperate, which correspond to the major global production environments. Reproductive phenology plays a key role in lentil adaptation to this diverse ecogeographic variation. Here, we dissect the characteristic earliness of the pilosae ecotype, suited to the typically short cropping season of South Asian environments. We identified two loci, DTF6a and DTF6b, at which dominant alleles confer early flowering, and we show that DTF6a alone is sufficient to confer early flowering under extremely short photoperiods. Genomic synteny confirmed the presence of a conserved cluster of three florigen (FT) gene orthologues among potential candidate genes, and expression analysis in near-isogenic material showed that the early allele is associated with a strong derepression of the FTa1 gene in particular. Sequence analysis revealed a 7.4 kb deletion in the FTa1-FTa2 intergenic region in the pilosae parent, and a wide survey of >350 accessions with diverse origin showed that the dtf6a allele is predominant in South Asian material. Collectively, these results contribute to understanding the molecular basis of global adaptation in lentil, and further emphasize the importance of this conserved genomic region for adaptation in temperate legumes generally.

The flowering hormone florigen accelerates secondary cell wall biogenesis to harmonize vascular maturation with reproductive development.

Florigen, a proteinaceous hormone, functions as a universal long-range promoter of flowering and concurrently as a generic growth-attenuating hormone across leaf and stem meristems. In flowering plants, the transition from the vegetative phase to the reproductive phase entails the orchestration of new growth coordinates and a global redistribution of resources, signals, and mechanical loads among organs. However, the ultimate cellular processes governing the adaptation of the shoot system to reproduction remain unknown. We hypothesized that if the mechanism for floral induction is universal, then the cellular metabolic mechanisms underlying the conditioning of the shoot system for reproduction would also be universal and may be best regulated by florigen itself. To understand the cellular basis for the vegetative functions of florigen, we explored the radial expansion of tomato stems. RNA-Seq and complementary genetic and histological studies revealed that florigen of endogenous, mobile, or induced origins accelerates the transcription network navigating secondary cell wall biogenesis as a unit, promoting vascular maturation and thereby adapting the shoot system to the developmental needs of the ensuing reproductive phase it had originally set into motion. We then demonstrated that a remarkably stable and broadly distributed florigen promotes MADS and MIF genes, which in turn regulate the rate of vascular maturation and radial expansion of stems irrespective of flowering or florigen level. The dual acceleration of flowering and vascular maturation by florigen provides a paradigm for coordinated regulation of independent global developmental programs.

Functional Conservation and Divergence of MOS1 That Controls Flowering Time and Seed Size in Rice and Arabidopsis.

The heading date and grain size are two essential traits affecting rice yield. Here, we found that OsMOS1 promotes rice heading and affects its grain size. Knocking out OsMOS1 delayed heading, while the overexpression of OsMOS1 promoted heading in rice under long-day conditions. The transcriptions of the heading activators Ehd1, Hd3a, and RFT1 were decreased and the heading repressor Hd1 was increased in the osmos1 mutant. Conversely, the overexpression of OsMOS1 promoted the expressions of Ehd1, Hd3a, and RFT1, but inhibited the expression of Hd1. This suggests that OsMOS1 may control heading in rice by modulating the transcriptions of Ehd1, Hd3a, RFT1, and Hd1. In addition, knocking out OsMOS1 led to larger grains with longer grain lengths and higher grain weights. The seed cell size measurement showed that the cell lengths and cell widths of the outer glume epidermal cells of the osmos1 mutant were greater than those of the wild type. Furthermore, we also found that the overexpression of OsMOS1 in the Arabidopsis mos1 mutant background could suppress its phenotypes of late flowering and increased seed size. Thus, our study shows a conserved function of MOS1 in rice and Arabidopsis, and these findings shed light on the heading and seed size regulation in rice and suggest that OsMOS1 is a promising target for rice yield improvement.

The florigen interactor BdES43 represses flowering in the model temperate grass Brachypodium distachyon.

FLOWERING LOCUS T (FT) protein, physiologically florigen, has been identified as a system integrator of numerous flowering time pathways in many studies, and its homologs are found throughout the plant lineage. It is important to uncover how precisely florigenic homologs contribute to flowering initiation and how these factors interact genetically. Here we dissected the function of Brachypodium FT orthologs BdFTL1 and BdFTL2 using overexpression and gene-editing experiments. Transgenic assays showed that both BdFTL1 and BdFTL2 could promote flowering, whereas BdFTL2 was essential for flowering initiation. Notably, BdFTL1 is subject to alternative splicing (AS), and its transcriptional level and AS are significantly affected by BdFTL2. Additionally, BdFTL2 could bind with the PHD-containing protein BdES43, an H3K4me3 reader. Furthermore, BdES43 was antagonistic to BdFTL2 in flowering initiation in a transcription-dependent manner and significantly affected BdFTL1 expression. BdFTL2, BdES43 and H3K4me3 also had highly similar distribution patterns within the BdFTL1 locus, indicating their interplay in regulating target genes. Taken together, florigen BdFTL2 functions as a potential epigenetic effector of BdFTL1 by interacting with a BdES43-H3K4me3 complex. This finding provides an additional insight for the regulatory mechanism underlying the multifaceted roles of florigen.

Components and Functional Diversification of Florigen Activation Complexes in Cotton.

In shoot apex cells of rice, a hexameric florigen activation complex (FAC), comprising flowering locus T (FT), 14-3-3 and the basic leucine zipper transcription factor FD, activates downstream target genes and regulates several developmental transitions, including flowering. The allotetraploid cotton (Gossypium hirsutum L.) contains only one FT locus in both of the A- and D-subgenomes. However, there is limited information regarding cotton FACs. Here, we identified a 14-3-3 protein that interacts strongly with GhFT in the cytoplasm and the nuclei, and five FD homoeologous gene pairs were characterized. In vivo, all five GhFD proteins interacted with Gh14-3-3 and GhFT in the nucleus. GhFT, 14-3-3 and all the GhFDs interacted in the nucleus as well, suggesting that they formed a ternary complex. Virus-induced silencing of GhFD1, -2 and -4 in cotton delayed flowering and inhibited the expression of floral meristem identity genes. Silencing GhFD3 strongly decreased lateral root formation, suggesting a function in lateral root development. GhFD overexpression in Arabidopsis and transcriptional activation assays suggested that FACs containing GhFD1 and GhFD2 function mainly in promoting flowering with partial functional redundancy. Moreover, GhFD3 was specifically expressed in lateral root meristems and dominantly activated the transcription of auxin response factor genes, such as ARF19. Thus, the diverse functions of FACs may depend on the recruited GhFD. Creating targeted genetic mutations in the florigen system using Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) genome editing may fine-tune flowering and improve plant architecture.

Inter-species functional compatibility of the Theobroma cacao and Arabidopsis FT orthologs: 90 million years of functional conservation of meristem identity genes.

BACKGROUND: In angiosperms the transition to flowering is controlled by a complex set of interacting networks integrating a range of developmental, physiological, and environmental factors optimizing transition time for maximal reproductive efficiency. The molecular mechanisms comprising these networks have been partially characterized and include both transcriptional and post-transcriptional regulatory pathways. Florigen, encoded by FLOWERING LOCUS T (FT) orthologs, is a conserved central integrator of several flowering time regulatory pathways. To characterize the molecular mechanisms involved in controlling cacao flowering time, we have characterized a cacao candidate florigen gene, TcFLOWERING LOCUS T (TcFT). Understanding how this conserved flowering time regulator affects cacao plant's transition to flowering could lead to strategies to accelerate cacao breeding. RESULTS: BLAST searches of cacao genome reference assemblies identified seven candidate members of the CENTRORADIALIS/TERMINAL FLOWER1/SELF PRUNING gene family including a single florigen candidate. cDNA encoding the predicted cacao florigen was cloned and functionally tested by transgenic genetic complementation in the Arabidopsis ft-10 mutant. Transgenic expression of the candidate TcFT cDNA in late flowering Arabidopsis ft-10 partially rescues the mutant to wild-type flowering time. Gene expression studies reveal that TcFT is spatially and temporally expressed in a manner similar to that found in Arabidopsis, specifically, TcFT mRNA is shown to be both developmentally and diurnally regulated in leaves and is most abundant in floral tissues. Finally, to test interspecies compatibility of florigens, we transformed cacao tissues with AtFT resulting in the remarkable formation of flowers in tissue culture. The morphology of these in vitro flowers is normal, and they produce pollen that germinates in vitro with high rates. CONCLUSION: We have identified the cacao CETS gene family, central to developmental regulation in angiosperms. The role of the cacao's single FT-like gene (TcFT) as a general regulator of determinate growth in cacao was demonstrated by functional complementation of Arabidopsis ft-10 late-flowering mutant and through gene expression analysis. In addition, overexpression of AtFT in cacao resulted in precocious flowering in cacao tissue culture demonstrating the highly conserved function of FT and the mechanisms controlling flowering in cacao.