A Soybean Deletion Mutant That Moderates the Repression of Flowering by Cool Temperatures.
ABSTRACT: Ambient growing temperature and photoperiod are major environmental stimuli that summer annual crops use to adjust their reproductive phenology so as to maximize yield. Variation in flowering time among soybean (Glycine max) cultivars results mainly from allelic diversity at loci that control photoperiod sensitivity and FLOWERING LOCUS T (FT) orthologs. However, variation in the thermal regulation of flowering and its underlying mechanisms are poorly understood. In this study, we identified a novel mutant (ef1) that confers altered thermal regulation of flowering in response to cool ambient temperatures. Mapping analysis with simple sequence repeat (SSR) markers located the mutation in the upper part of chromosome 19, where no QTL for flowering has been previously reported. Fine-mapping and re-sequencing revealed that the mutation was caused by deletion of a 214 kbp genomic region that contains 11 annotated genes, including CONSTANS-LIKE 2b (COL2b), a soybean ortholog of Arabidopsis CONSTANS. Comparison of flowering times under different photo-thermal conditions revealed that early flowering in the mutant lines was most distinct under cool ambient temperatures. The expression of two FT orthologs, FT2a and FT5a, was dramatically downregulated by cool temperature, but the magnitude of the downregulation was lower in the mutant lines. Cool temperatures upregulated COL2b expression or delayed peak expression, particularly at the fourth trifoliate-leaf stage. Intriguingly, they also upregulated E1, a soybean-specific repressor of FT orthologs. Our results suggest that the ef1 mutation is involved in thermal regulation of flowering in response to cool ambient temperature, and the lack of COL2b in the mutant likely alleviates the repression of flowering by cool temperature. The ef1 mutant can be used as a novel gene resource in breeding soybean cultivars adapted to cool climate and in research to improve our understanding of thermal regulation of flowering in soybean.
Project description:BACKGROUND: Sorghum genotypes used for grain production in temperate regions are photoperiod insensitive and flower early avoiding adverse environments during the reproductive phase. In contrast, energy sorghum hybrids are highly photoperiod sensitive with extended vegetative phases in long days, resulting in enhanced biomass accumulation. SbPRR37 and SbGHD7 contribute to photoperiod sensitivity in sorghum by repressing expression of SbEHD1 and FT-like genes, thereby delaying flowering in long days with minimal influence in short days (PNAS_108:16469-16474, 2011; Plant Genome_in press, 2014). The GIGANTEA (GI)-CONSTANS (CO)-FLOWERING LOCUS T (FT) pathway regulates flowering time in Arabidopsis and the grasses (J Exp Bot_62:2453-2463, 2011). In long day flowering plants, such as Arabidopsis and barley, CONSTANS activates FT expression and flowering in long days. In rice, a short day flowering plant, Hd1, the ortholog of CONSTANS, activates flowering in short days and represses flowering in long days. RESULTS: Quantitative trait loci (QTL) that modify flowering time in sorghum were identified by screening Recombinant Inbred Lines (RILs) derived from BTx642 and Tx7000 in long days, short days, and under field conditions. Analysis of the flowering time QTL on SBI-10 revealed that BTx642 encodes a recessive CONSTANS allele containing a His106Tyr substitution in B-box 2 known to inactivate CONSTANS in Arabidopsis thaliana. Genetic analysis characterized sorghum CONSTANS as a floral activator that promotes flowering by inducing the expression of EARLY HEADING DATE 1 (SbEHD1) and sorghum orthologs of the maize FT genes ZCN8 (SbCN8) and ZCN12 (SbCN12). The floral repressor PSEUDORESPONSE REGULATOR PROTEIN 37 (PRR37) inhibits sorghum CONSTANS activity and flowering in long days. CONCLUSION: Sorghum CONSTANS is an activator of flowering that is repressed post-transcriptionally in long days by the floral inhibitor PRR37, contributing to photoperiod sensitive flowering in Sorghum bicolor, a short day plant.
Project description:BACKGROUND: CO and FT orthologs, belonging to the BBX and PEBP family, respectively, have important and conserved roles in the photoperiod regulation of flowering time in plants. Soybean genome experienced at least three rounds of whole genome duplications (WGDs), which resulted in multiple copies of about 75% of genes. Subsequent subfunctionalization is the main fate for paralogous gene pairs during the evolutionary process. RESULTS: The phylogenic relationships revealed that CO orthologs were widespread in the plant kingdom while FT orthologs were present only in angiosperms. Twenty-eight CO homologous genes and twenty-four FT homologous genes were gained in the soybean genome. Based on the collinear relationship, the soybean ancestral CO ortholog experienced three WGD events, but only two paralogous gene pairs (GmCOL1/2 and GmCOL5/13) survived in the modern soybean. The paralogous gene pairs, GmCOL1/2 or GmCOL5/13, showed similar expression patterns in pair but different between pairs, indicating that they functionally diverged. GmFTL1 to 7 were derived from the same ancestor prior to the whole genome triplication (WGT) event, and after the Legume WGD event the ancestor diverged into two branches, GmFTL3/5/7 and GmFTL1/2/4/6. GmFTL7 were truncated in the N-terminus compared to other FT-lineage genes, but ubiquitously expressed. Expressions of GmFTL1 to 6 were higher in leaves at the flowering stage than that at the seedling stage. GmFTL3 was expressed at the highest level in all tissues except roots at the seedling stage, and its circadian pattern was different from the other five ones. The transcript of GmFTL6 was highly accumulated in seedling roots. The circadian rhythms of GmCOL5/13 and GmFT1/2/4/5/6 were synchronized in a day, demonstrating the complicate relationship of CO-FT regulons in soybean leaves. Over-expression of GmCOL2 did not rescue the flowering phenotype of the Arabidopsis co mutant. However, ectopic expression of GmCOL5 did rescue the co mutant phenotype. All GmFTL1 to 6 showed flower-promoting activities in Arabidopsis. CONCLUSIONS: After three recent rounds of whole genome duplications in the soybean, the paralogous genes of CO-FT regulons showed subfunctionalization through expression divergence. Then, only GmCOL5/13 kept flowering-promoting activities, while GmFTL1 to 6 contributed to flowering control. Additionally, GmCOL5/13 and GmFT1/2/3/4/5/6 showed similar circadian expression profiles. Therefore, our results suggested that GmCOL5/13 and GmFT1/2/3/4/5/6 formed the complicate CO-FT regulons in the photoperiod regulation of flowering time in soybean.
Project description:Phytochromes confer the photoperiodic control of flowering in rice (Oryza sativa), a short-day plant. To better understand the molecular mechanisms of day-length recognition, we examined the interaction between phytochrome signals and circadian clocks in photoperiodic-flowering mutants of rice. Monitoring behaviors of circadian clocks revealed that phase setting of circadian clocks is not affected either under short-day (SD) or under long-day (LD) conditions in a phytochrome-deficient mutant that shows an early-flowering phenotype with no photoperiodic response. Non-24-hr-light/dark-cycle experiments revealed that a rice counterpart gene of Arabidopsis CONSTANS (CO), named PHOTOPERIOD SENSITIVITY 1 (Heading date 1) [SE1 (Hd1)], functions as an output of circadian clocks. In addition, the phytochrome deficiency does not affect the diurnal mRNA expression of SE1 upon floral transition. Downstream floral switch genes were further identified with rice orthologs of Arabidopsis FLOWERING LOCUS T (FT). Our RT-PCR data indicate that phytochrome signals repress mRNA expression of FT orthologs, whereas SE1 can function to promote and suppress mRNA expression of the FT orthologs under SD and LD, respectively. This SE1 transcriptional activity may be posttranscriptionally regulated and may depend on the coincidence with Pfr phytochromes. We propose a model to explain how a short-day plant recognizes the day length in photoperiodic flowering.
Project description:A soybean MADS box gene GmGAL2 (Glycine max AGAMOUS Like 2), a homolog of AGL11/STK, was investigated in transgenic Arabidopsis lines. Ectopic expression of GmGAL2 in Arabidopsis enhanced flowering, under both long-day and short-day conditions, by promoting expression of key flowering genes, CONSTANS (CO) and FLOWERING LOCUS T (FT), and lowering expression of floral inhibiter FLOWERING LOCUS C (FLC). Moreover, frequency of silique pod set was also lower in transgenic compared to control Arabidopsis plants. RT-PCR results revealed that GmGAL2 was primarily expressed in the flowers and pods of soybean plants, GmGAL2 expressed higher in SD than LD in soybean.
Project description:Many plants are incapable of flowering in inductive daylengths during the early juvenile vegetative phase (JVP). Arabidopsis mutants with reduced expression of TEMPRANILLO (TEM), a repressor of flowering locus T (FT) had a shorter JVP than wild-type plants. Reciprocal changes in mRNA expression of TEM and FT were observed in both Arabidopsis and antirrhinum, which correlated with the length of the JVP. FT expression was induced just prior to the end of the JVP and levels of TEM1 mRNA declined rapidly at the time when FT mRNA levels were shown to increase. TEM orthologs were isolated from antirrhinum (AmTEM) and olive (OeTEM) and were expressed most highly during their juvenile phase. AmTEM functionally complemented AtTEM1 in the tem1 mutant and over-expression of AmTEM prolonged the JVP through repression of FT and CONSTANS (CO). We propose that TEM may have a general role in regulating JVP in herbaceous and woody species.
Project description:Day length and ambient temperature are major stimuli controlling flowering time. To understand flowering mechanisms in more natural conditions, we explored the effect of daily light and temperature changes on Arabidopsis thaliana. Seedlings were exposed to different day/night temperature and day-length treatments to assess expression changes in flowering genes. Cooler temperature treatments increased CONSTANS (CO) transcript levels at night. Night-time CO induction was diminished in flowering bhlh (fbh)-quadruple mutants. FLOWERING LOCUS T (FT) transcript levels were reduced at dusk, but increased at the end of cooler nights. The dusk suppression, which was alleviated in short vegetative phase (svp) mutants, occurred particularly in younger seedlings, whereas the increase during the night continued over 2 wk. Cooler temperature treatments altered the levels of FLOWERING LOCUS M-? (FLM-?) and FLM-? splice variants. FT levels correlated strongly with flowering time across treatments. Day/night temperature changes modulate photoperiodic flowering by changing FT accumulation patterns. Cooler night-time temperatures enhance FLOWERING BHLH (FBH)-dependent induction of CO and consequently increase CO protein. When plants are young, cooler temperatures suppress FT at dusk through SHORT VEGETATIVE PHASE (SVP) function, perhaps to suppress precocious flowering. Our results suggest day length and diurnal temperature changes combine to modulate FT and flowering time.
Project description:Breeding vegetative crops (e.g. beets, cabbage, forage grasses) is challenged by two conflicting aims. For field production, flowering must be avoided while flowering and seed set is necessary for breeding and seed production. The biennial species sugar beet makes shoot elongation ('bolting') followed by flowering after a long period of cold temperatures. Field production in northern geographical regions starts in spring. A thickened storage root is formed only during vegetative growth. It is expected that winter beets, which are sown before winter would have a much higher yield potential. However, field production was not possible so far due to bolting after winter. We propose a strategy to breed winter beets exploiting haplotype variation at two major bolting time loci, B and B2. Both genes encode transcription factors controlling the expression of two orthologs of the Arabidopsis gene FLOWERING LOCUS T (FT). We detected an epistatic interaction between both genes because F<sub>2</sub> plants homozygous for two B/B2 mutant alleles did not bolt even after vernalization. Fluorescence complementation studies revealed that both proteins form a heterodimer in vivo. In non-bolting plants, the bolting activator BvFT2 was completely downregulated whereas the repressor BvFT1 was upregulated which suggests that both genes acquire a CONSTANS (CO) like function in beet. Like CO, B and B2 proteins house CCT and BBX domains which, in contrast to CO are split between the two beet genes. We propose an alternative regulation of FT orthologs in beet that can be exploited to breed winter beets.
Project description:The timing of both flowering and maturation determine crop adaptability and productivity. Soybean (Glycine max) is cultivated across a wide range of latitudes. The molecular-genetic mechanisms for flowering in soybean have been determined for photoperiodic responses to long days (LDs), but remain only partially determined for the delay of flowering under short-day conditions, an adaptive trait of cultivars grown in lower latitudes. Here, we characterized the late-flowering (LF) habit introduced from the Thai cultivar K3 into a photoperiod-insensitive genetic background under different photo-thermal conditions, and we analyzed the genetic basis using quantitative trait locus (QTL) mapping. The LF habit resulted from a basic difference in the floral induction activity and from the suppression of flowering, which was caused by red light-enriched LD lengths and higher temperatures, during which FLOWERING LOCUS T (FT) orthologs, FT2a and FT5a, were strongly down-regulated. QTL mapping using gene-specific markers for flowering genes E2, FT2a and FT5a and 829 single nucleotide polymorphisms obtained from restriction-site associated DNA sequencing detected three QTLs controlling the LF habit. Of these, a QTL harboring FT2a exhibited large and stable effects under all the conditions tested. A resequencing analysis detected a nonsynonymous substitution in exon 4 of FT2a from K3, which converted the glycine conserved in FT-like proteins to the aspartic acid conserved in TERMINAL FLOWER 1-like proteins (floral repressors), suggesting a functional depression in the FT2a protein from K3. The effects of the remaining two QTLs, likely corresponding to E2 and FT5a, were environment dependent. Thus, the LF habit from K3 may be caused by the functional depression of FT2a and the down-regulation of two FT genes by red light-enriched LD conditions and high temperatures.
Project description:Appropriate control of flowering time is crucial for crop yield and the reproductive success of plants. Flowering can be induced by a number of molecular pathways that respond to internal and external signals. In Arabidopsis, expression of the key florigenic signal FLOWERING LOCUS T (FT) is positively regulated by CONSTANS (CO) a BBX protein sharing high sequence similarity with 16 CO-like proteins. Within this study, we investigated the role of the Arabidopsis CONSTANS-LIKE 4 (COL4), whose role in flowering control was unknown. We demonstrate that, unlike CO, COL4 is a flowering repressor in long days (LD) and short days (SD) and acts on the expression of FT and FT-like genes as well as on SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1). Reduction of COL4 expression level leads to an increase of FT and APETALA 1 (AP1) expression and to accelerated flowering, while the increase of COL4 expression causes a flowering delay. Further, the observed co-localization of COL4 protein and CO in nuclear speckles supports the idea that the two act as an antagonistic pair of transcription factors. This interaction may serve the fine-tuning of flowering time control and other light dependent plant developmental processes.
Project description:The zinc finger transcription factor CONSTANS has a well-established central role in the mechanism for photoperiod sensing in Arabidopsis, integrating light and circadian clock signals to upregulate the florigen gene FT under long-day but not short-day conditions. Although CONSTANS-LIKE (COL) genes in other species have also been shown to regulate flowering time, it is not clear how widely this central role in photoperiod sensing is conserved. Legumes are a major plant group and various legume species show significant natural variation for photoperiod responsive flowering. Orthologs of several Arabidopsis genes have been shown to participate in photoperiodic flowering in legumes, but the possible function of COL genes as integrators of the photoperiod response has not yet been examined in detail. Here we characterize the COL family in the temperate long-day legume Medicago truncatula, using expression analyses, reverse genetics, transient activation assays and Arabidopsis transformation. Our results provide several lines of evidence suggesting that COL genes are unlikely to have a central role in the photoperiod response mechanism in this species.