Diurnal Expression Pattern, Allelic Variation, and Association Analysis Reveal Functional Features of the E1 Gene in Control of Photoperiodic Flowering in Soybean.
ABSTRACT: Although four maturity genes, E1 to E4, in soybean have been successfully cloned, their functional mechanisms and the regulatory network of photoperiodic flowering remain to be elucidated. In this study, we investigated how the diurnal expression pattern of the E1 gene is related to photoperiodic length; and to what extent allelic variation in the B3-like domain of the E1 gene is associated with flowering time phenotype. The bimodal expression of the E1 gene peaked first at around 2 hours after dawn in long-day condition. The basal expression level of E1 was enhanced by the long light phase, and decreased by duration of dark. We identified a 5bp (3 SNP and 2-bp deletion) mutation, referred to an e1-b3a, which occurs in the middle of B3 domain of the E1 gene in the early flowering cultivar Yanhuang 3. Subcellular localization analysis showed that the putative truncated e1-b3a protein was predominately distributed in nuclei, indicating the distribution pattern of e1-b3a was similar to that of E1, but not to that of e1-as. Furthermore, genetic analysis demonstrated allelic variations at the E1 locus significantly underlay flowering time in three F2 populations. Taken together, we can conclude the legume specific E1 gene confers some special features in photoperiodic control of flowering in soybean. Further characterization of the E1 gene will extend our understanding of the soybean flowering pathway in soybean.
Project description:Gene regulatory networks involved in flowering time and photoperiodic responses in legumes remain unknown. Although the major maturity gene E1 has been successfully deciphered in soybean, knowledge on the functional conservation of this gene is limited to a certain extent to E1 homologs in legumes. The ectopic expression of Phvul.009G204600 (PvE1L), an E1 homolog from common bean, delayed the onset of flowering in soybean. By contrast, the ectopic expression of Medtr2g058520 (MtE1L) from Medicago truncatula did not affect the flowering of soybean. Characterization of the late-flowering mte1l mutant indicated that MtE1L promoted flowering in Medicago truncatula. Moreover, all transgenic E1, PvE1L and MtE1L soybean lines exhibited phenotypic changes in terms of plant height. Transgenic E1 or PvE1L plants were taller than the wild-type, whereas transgenic MtE1L plants produced dwarf phenotype with few nodes and short internode. Thus, functional conservation and diversification of E1 family genes from legumes in the regulation of flowering and plant growth may be associated with lineage specification and genomic duplication.
Project description:Photoperiodic flowering, a plant response to seasonal photoperiod changes in the control of reproductive transition, is an important agronomic trait that has been a central target of crop domestication and modern breeding programs. However, our understanding about the molecular mechanisms of photoperiodic flowering regulation in crop species is lagging behind. To better understand the regulatory gene networks controlling photoperiodic flowering of soybeans, we elucidated global gene expression patterns under different photoperiod regimes using the near isogenic lines (NILs) of maturity loci (E loci). Transcriptome signatures identified the unique roles of the E loci in photoperiodic flowering and a set of genes controlled by these loci. To elucidate the regulatory gene networks underlying photoperiodic flowering regulation, we developed the network inference algorithmic package CausNet that integrates sparse linear regression and Granger causality heuristics, with Gaussian approximation of bootstrapping to provide reliability scores for predicted regulatory interactions. Using the transcriptome data, CausNet inferred regulatory interactions among soybean flowering genes. Published reports in the literature provided empirical verification for several of CausNet's inferred regulatory interactions. We further confirmed the inferred regulatory roles of the flowering suppressors GmCOL1a and GmCOL1b using GmCOL1 RNAi transgenic soybean plants. Combinations of the alleles of GmCOL1 and the major maturity locus E1 demonstrated positive interaction between these genes, leading to enhanced suppression of flowering transition. Our work provides novel insights and testable hypotheses in the complex molecular mechanisms of photoperiodic flowering control in soybean and lays a framework for de novo prediction of biological networks controlling important agronomic traits in crops.
Project description:The complex and coordinated regulation of flowering has high ecological and agricultural significance. The maturity locus E1 has a large impact on flowering time in soybean, but the molecular basis for the E1 locus is largely unknown. Through positional cloning, we delimited the E1 locus to a 17.4-kb region containing an intron-free gene (E1). The E1 protein contains a putative bipartite nuclear localization signal and a region distantly related to B3 domain. In the recessive allele, a nonsynonymous substitution occurred in the putative nuclear localization signal, leading to the loss of localization specificity of the E1 protein and earlier flowering. The early-flowering phenotype was consistently observed in three ethylmethanesulfonate-induced mutants and two natural mutations that harbored a premature stop codon or a deletion of the entire E1 gene. E1 expression was significantly suppressed under short-day conditions and showed a bimodal diurnal pattern under long-day conditions, suggesting its response to photoperiod and its dominant effect induced by long day length. When a functional E1 gene was transformed into the early-flowering cultivar Kariyutaka with low E1 expression, transgenic plants carrying exogenous E1 displayed late flowering. Furthermore, the transcript abundance of E1 was negatively correlated with that of GmFT2a and GmFT5a, homologues of FLOWERING LOCUS T that promote flowering. These findings demonstrated the key role of E1 in repressing flowering and delaying maturity in soybean. The molecular identification of the maturity locus E1 will contribute to our understanding of the molecular mechanisms by which a short-day plant regulates flowering time and maturity.
Project description:The time to flowering and maturity are ecologically and agronomically important traits for soybean landrace and cultivar adaptation. As a typical short-day crop, long day conditions in the high-latitude regions require soybean cultivars with photoperiod insensitivity that can mature before frost. Although the molecular basis of four major E loci (E1 to E4) have been deciphered, it is not quite clear whether, or to what degree, genetic variation and the expression level of the four E genes are associated with the time to flowering and maturity of soybean cultivars. In this study, we genotyped 180 cultivars at E1 to E4 genes, meanwhile, the time to flowering and maturity of those cultivars were investigated at six geographic locations in China from 2011 to 2012 and further confirmed in 2013. The percentages of recessive alleles at E1, E2, E3 and E4 loci were 38.34%, 84.45%, 36.33%, and 7.20%, respectively. Statistical analysis showed that allelic variations at each of four loci had a significant effect on flowering time as well as maturity. We classified the 180 cultivars into eight genotypic groups based on allelic variations of the four major E loci. The genetic group of e1-nf representing dysfunctional alleles at the E1 locus flowered earliest in all the geographic locations. In contrast, cultivars in the E1E2E3E4 group originated from the southern areas flowered very late or did not flower before frost at high latitude locations. The transcriptional abundance of functional E1 gene was significantly associated with flowering time. However, the ranges of time to flowering and maturity were quite large within some genotypic groups, implying the presence of some other unknown genetic factors that are involved in control of flowering time or maturity. Known genes (e.g. E3 and E4) and other unknown factors may function, at least partially, through regulation of the expression of the E1 gene.
Project description:BACKGROUND: Absence of or low sensitivity to photoperiod is necessary for short-day crops, such as rice and soybean, to adapt to high latitudes. Photoperiod insensitivity in soybeans is controlled by two genetic systems and involves three important maturity genes: E1, a repressor for two soybean orthologs of Arabidopsis FLOWERING LOCUS T (GmFT2a and GmFT5a), and E3 and E4, which are phytochrome A genes. To elucidate the diverse mechanisms underlying photoperiod insensitivity in soybean, we assessed the genotypes of four maturity genes (E1 through E4) in early-flowering photoperiod-insensitive cultivars and their association with post-flowering responses. RESULTS: We found two novel dysfunctional alleles in accessions originally considered to have a dominant E3 allele according to known DNA markers. The E3 locus, together with E1 and E4, contained multiple dysfunctional alleles. We identified 15 multi-locus genotypes, which we subdivided into 6 genotypic groups by classifying their alleles by function. Of these, the e1-as/e3/E4 genotypic group required an additional novel gene (different from E1, E3, and E4) to condition photoperiod insensitivity. Despite their common pre-flowering photoperiod insensitivity, accessions with different multi-locus genotypes responded differently to the post-flowering photoperiod. Cultivars carrying E3 or E4 were sensitive to photoperiod for post-flowering characteristics, such as reproductive period and stem growth after flowering. The phytochrome A-regulated expression of the determinate growth habit gene Dt1, an ortholog of Arabidopsis TERMINAL FLOWER1, was involved in the persistence of the vegetative activity at the stem apical meristem of flower-induced plants under long-day conditions. CONCLUSIONS: Diverse genetic mechanisms underlie photoperiod insensitivity in soybean. At least three multi-locus genotypes consisting of various allelic combinations at E1, E3, and E4 conferred pre-flowering photoperiod insensitivity to soybean cultivars but led to different responses to photoperiod during post-flowering vegetative and reproductive development. The phyA genes E3 and E4 are major controllers underlying not only pre-flowering but also post-flowering photoperiod responses. The current findings improve our understanding of genetic diversity in pre-flowering photoperiod insensitivity and mechanisms of post-flowering photoperiod responses in soybean.
Project description:CONSTANS (CO) plays a central role in photoperiodic flowering control of plants. However, much remains unknown about the function of the CO gene family in soybean and the molecular mechanisms underlying short-day photoperiodic flowering of soybean. We identified 26 CO homologs (GmCOLs) in the soybean genome, many of them previously unreported. Phylogenic analysis classified GmCOLs into three clades conserved among flowering plants. Two homeologous pairs in Clade I, GmCOL1a/GmCOL1b and GmCOL2a/GmCOL2b, showed the highest sequence similarity to Arabidopsis CO. The mRNA abundance of GmCOL1a and GmCOL1b exhibited a strong diurnal rhythm under flowering-inductive short days and peaked at dawn, which coincided with the rise of GmFT5a expression. In contrast, the mRNA abundance of GmCOL2a and GmCOL2b was extremely low. Our transgenic study demonstrated that GmCOL1a, GmCOL1b, GmCOL2a and GmCOL2b fully complemented the late flowering effect of the co-1 mutant in Arabidopsis. Together, these results indicate that GmCOL1a and GmCOL1b are potential inducers of flowering in soybean. Our data also indicate rapid regulatory divergence between GmCOL1a/GmCOL1b and GmCOL2a/GmCOL2b but conservation of their protein function. Dynamic evolution of GmCOL regulatory mechanisms may underlie the evolution of photoperiodic signaling in soybean.
Project description:Photoperiodic control of flowering time is believed to affect latitudinal distribution of plants. The blue light receptor CRY2 regulates photoperiodic flowering in the experimental model plant Arabidopsis thaliana. However, it is unclear whether genetic variations affecting cryptochrome activity or expression is broadly associated with latitudinal distribution of plants. We report here an investigation of the function and expression of two cryptochromes in soybean, GmCRY1a and GmCRY2a. Soybean is a short-day (SD) crop commonly cultivated according to the photoperiodic sensitivity of cultivars. Both cultivated soybean (Glycine max) and its wild relative (G. soja) exhibit a strong latitudinal cline in photoperiodic flowering. Similar to their Arabidopsis counterparts, both GmCRY1a and GmCRY2a affected blue light inhibition of cell elongation, but only GmCRY2a underwent blue light- and 26S proteasome-dependent degradation. However, in contrast to Arabidopsis cryptochromes, soybean GmCRY1a, but not GmCRY2a, exhibited a strong activity promoting floral initiation, and the level of protein expression of GmCRY1a, but not GmCRY2a, oscillated with a circadian rhythm that has different phase characteristics in different photoperiods. Consistent with the hypothesis that GmCRY1a is a major regulator of photoperiodic flowering in soybean, the photoperiod-dependent circadian rhythmic expression of the GmCRY1a protein correlates with photoperiodic flowering and latitudinal distribution of soybean cultivars. We propose that genes affecting protein expression of the GmCRY1a protein play an important role in determining latitudinal distribution of soybeans.
Project description:BACKGROUND AND AIMS: The timing of flowering has a direct impact on successful seed production in plants. Flowering of soybean (Glycine max) is controlled by several E loci, and previous studies identified the genes responsible for the flowering loci E1, E2, E3 and E4. However, natural variation in these genes has not been fully elucidated. The aims of this study were the identification of new alleles, establishment of allele diagnoses, examination of allelic combinations for adaptability, and analysis of the integrated effect of these loci on flowering. METHODS: The sequences of these genes and their flanking regions were determined for 39 accessions by primer walking. Systematic discrimination among alleles was performed using DNA markers. Genotypes at the E1-E4 loci were determined for 63 accessions covering several ecological types using DNA markers and sequencing, and flowering times of these accessions at three sowing times were recorded. KEY RESULTS: A new allele with an insertion of a long interspersed nuclear element (LINE) at the promoter of the E1 locus (e1-re) was identified. Insertion and deletion of 36 bases in the eighth intron (E2-in and E2-dl) were observed at the E2 locus. Systematic discrimination among the alleles at the E1-E3 loci was achieved using PCR-based markers. Allelic combinations at the E1-E4 loci were found to be associated with ecological types, and about 62-66 % of variation of flowering time could be attributed to these loci. CONCLUSIONS: The study advances understanding of the combined roles of the E1-E4 loci in flowering and geographic adaptation, and suggests the existence of unidentified genes for flowering in soybean.
Project description:BACKGROUND:Soybean is native to the temperate zones of East Asia. Poor yields of soybean in West African countries may be partially attributed to inadequate adaptation of soybean to tropical environments. Adaptation will require knowledge of the effects of allelic combinations of major maturity genes (E1, E2, and E3) and stem architecture. The long juvenile trait (J) influences soybean flowering time in short, ~?12?h?days, which characterize tropical latitudes. Soybean plant architecture includes determinate or indeterminate stem phenotypes controlled by the Dt1 gene. Understanding the influence of these genetic components on plant development and adaptation is key to optimize phenology and improve soybean yield potential in tropical environments. RESULTS:Soybean lines from five recombinant inbred populations were developed that varied in their combinations of targeted genes. The soybean lines were field tested in multiple environments and characterized for days to flowering (DTF), days to maturity (DTM), and plant height in locations throughout northern Ghana, and allelic combinations were determined for each line for associating genotype with phenotype. The results revealed significant differences based on genotype for DTF and DTM and allowed the comparison of different variant alleles of those genes. The mutant alleles of J and E1 had significant impact on DTF and DTM, and alleles of those genes interacted with each other for DTF but not DTM. The Dt1 gene significantly influenced plant height but not DTF or DTM. CONCLUSIONS:This research identified major and minor effect alleles of soybean genes that can be combined to control DTF, DTM, and plant height in short day tropical environments in Ghana. These phenotypes contribute to adaptation to a low latitude environment that can be optimized in a soybean breeding program with targeted selection of desired allele combinations. The knowledge of the genetic control of these traits will enhance molecular breeding to produce optimally adapted soybean varieties targeted to tropical environments.
Project description:Soybean time of flowering and maturity are genetically controlled by E genes. Different allelic combinations of these genes determine soybean adaptation to a specific latitude. The paper describes the first attempt to assess adaptation of soybean genotypes developed and realized at Institute of Field and Vegetable Crops, Novi Sad, Serbia [Novi Sad (NS) varieties and breeding lines] based on E gene variation, as well as to comparatively assess E gene variation in North-American (NA), Chinese, and European genotypes, as most of the studies published so far deal with North-American and Chinese cultivars and breeding material. Allelic variation and distribution of the major maturity genes (E1, E2, E3, and E4) has been determined in 445 genotypes from soybean collections of NA ancestral lines, Chinese germplasm, and European varieties, as well as NS varieties and breeding lines. The study showed that allelic combinations of E1-E4 genes significantly determined the adaptation of varieties to different geographical regions, although they have different impacts on maturity. In general, each collection had one major E genotype haplogroup, comprising over 50% of the lines. The exceptions were European varieties that had two predominant haplogroups and NA ancestral lines distributed almost evenly among several haplogroups. As e1-as/e2/E3/E4 was the most common genotype in NS population, present in the best-performing genotypes in terms of yield, this specific allele combination was proposed as the optimal combination for the environments of Central-Eastern Europe.