A loss-of-function mutant allele of a glycosyl hydrolase gene has been co-opted for seed weight control during soybean domestication

The resultant DNA from loss-of-function mutation can be recruited in biological evolution and development. Here, we present such a rare and potential case of “to gain by loss” as a neomorphic mutation during soybean domestication for increasing seed weight. Using a population derived from a chromosome segment substitution line of Glycine max(SN14) and Glycine soja(ZYD06), a quantitative trait locus(QTL) of 100-seed weight(q HSW) was mapped on chromosome 11, corresponding to a truncated β-1, 3-gl...     

Metabolic engineering of soybean affords improved phytosterol seed traits

Different combinations of three rate‐limiting enzymes in phytosterol biosynthesis, the Arabidopsis thaliana hydroxyl methylglutaryl CoA1 (HMGR1) catalytic subunit linked to either constitutive or seed‐specific β‐conglycinin promoter, and the Glycine max sterol methyltransferase1 (SMT1) and sterol methyltransferase2‐2 (SMT2‐2) genes, under the control of seed‐specific Glycinin‐1 and Beta‐phaseolin promoters, respectively, were engineered in soybean plants. Mature seeds of transgenic plants displayed modest increases in total sterol content, which points towards a tight control of phytosterol biosynthesis. However, in contrast to wild‐type seeds that accumulated about 35% of the total sterol in the form of intermediates, in the engineered seeds driven by a seed‐specific promoter, metabolic flux was directed to Δ⁵‐24‐alkyl sterol formation (99% of total sterol). The engineered effect of end‐product sterol (sitosterol, campesterol, and stigmasterol) over‐production in soybean seeds resulted in an approximately 30% increase in overall sitosterol synthesis, a desirable trait for oilseeds and human health. In contradistinction, increased accumulation of cycloartenol and 24(28)‐methylencylartanol (55% of the total sterol) was detected in plants harbouring the constitutive t‐HMGR1 gene, consistent with the previous studies. Our results support the possibility that metabolic flux of the phytosterol family pathway is differentially regulated in leaves and seeds.     

CLUSTERED PRIMARY BRANCH 1, a new allele of DWARF11, controls panicle architecture and seed size in rice

Panicle architecture and seed size are important agronomic traits that directly determine grain yield in rice (Oryza sativa L.). Although a number of key genes controlling panicle architecture and seed size have been cloned and characterized in recent years, their genetic and molecular mechanisms remain unclear. In this study, we identified a mutant that produced panicles with fascicled primary branching and reduced seeds in size. We isolated the underlying CLUSTERED PRIMARY BRANCH 1 (CPB1) gene, a new allele of DWARF11 (D11) encoding a cytochrome P450 protein involved in brassinosteroid (BR) biosynthesis pathway. Genetic transformation experiments confirmed that a His360Leu amino acid substitution residing in the highly conserved region of CPB1/D11 was responsible for the panicle architecture and seed size changes in the cpb1 mutants. Overexpression of CPB1/D11 under the background of cpb1 mutant not only rescued normal panicle architecture and plant height, but also had a larger leaf angle and seed size than the controls. Furthermore, the CPB1/D11 transgenic plants driven by panicle‐specific promoters can enlarge seed size and enhance grain yield without affecting other favourable agronomic traits. These results demonstrated that the specific mutation in CPB1/D11 influenced development of panicle architecture and seed size, and manipulation of CPB1/D11 expression using the panicle‐specific promoter could be used to increase seed size, leading to grain yield improvement in rice.     

Seed traits and bird species influence the dispersal parameters of wetland plants

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Seed weight and germination behavior of the submerged plant Potamogeton pectinatus in the arid zone of northwest China

Variation in seed weight is common within and among plant species, but few studies have attempted to document the pattern of seed weight and germination attributes for aquatic macrophytes at a large scale. This study examined within‐species variation in seed weight and germination attributes and the effects of environmental factors on seed traits of the submerged plant Potamogeton pectinatus in the arid zone of northwest China. Our results showed that the average seed weight was 0.24 g per 100 seeds with a coefficient of variation (CV) of 28.4% among the eight P. pectinatus populations. The total germination fraction of seeds of P. pectinatus was relatively poor, less than 35% in seven P. pectinatus populations, and the lowest germination percentage found was only 2%. There were significant differences in seed weight, time to onset of germination, and total germination fraction among the eight different populations. Hierarchical partitioning analysis showed a strongly positive correlation between seed weight and water temperature and pH. Seed weight and the maternal environmental factors significantly affected both time to initiation of germination and total germination fraction. Our results suggest that (1) seed weight variation in P. pectinatus primarily is the result of temperature variation during fruit development; (2) relatively poor germination fraction suggests that seeds are relatively unimportant in the short‐term survival of populations and that it may be another adaptive trait allowing plants to take place in the right place and at the right time, especially in harsh environment; and (3) variation in seed germination traits should be determined by local environmental and intrinsic factors that interact in a complex fashion.     

Promoter regions of cysteine endopeptidase genes from legumes confer germination-specific expression in transgenic tobacco seeds

Cysteine endopeptidases, SH-EP from Vigna mungo and EP-C1 from Phaseolus vulgaris, act to degrade seed storage protein during seed germination. Using transgenic tobacco plants, expression of SH-EP and promoter activity of the EP-C1 gene were analyzed in transgenic tobacco plants. The promoters of the two genes in tobacco seeds showed germination-specific activation, although post-translational processing of SH-EP and regulatory regions of promoter of the gene for EP-C1 were found to differ between leguminous seeds and transgenic tobacco seeds.     

Increasing maize seed weight by enhancing the cytoplasmic ADP-glucose pyrophosphorylase activity in transgenic maize plants

ADP-glucose pyrophosphorylase (AGPase) plays a key role in regulating starch biosynthesis in cereal seeds and is likely the most important determinant of seed strength. The Escherichia coli mutant glgC gene (glgC16), which encodes a highly active and allosterically insensitive AGPase, was introduced into maize (Zea mays L.) under the control of an endosperm-specific promoter. Developing seeds from transgenic maize plants showed up to 2–4-fold higher levels of AGPase activity in the presence of 5 mM inorganic phosphate (Pi). Transgenic plants with higher cytoplasmic AGPase activity under Pi-inhibitory conditions showed increases (13–25%) in seed weight over the untransformed control. In addition, in all transgenic maize plants, the seeds were fully filled, and the seed number of transgenic plants had no significant difference compared with that of untransformed control. These results indicate that increasing cytoplasmic AGPase activity has a marked effect on sink activity and, in turn, seed weight in transgenic maize plants.     

Growth,yield and reproduction of dwarf tomato grown under simulated microgravity conditions

Abstract

A closed hydroponic system combined with a horizontal uniaxial clinostat has been used to grow tomato plants (Solanum lycopersicum L.) under simulated microgravity conditions. The study was carried out to evaluate the quanti-qualitative traits (growth, yield and quality) of the dwarf tomato variety ‘Micro-Tom’ grown under simulated microgravity conditions and to determine if tomato plants would complete their life cycle (‘seed-to-seed’). Morphological and growth characteristics of ‘Micro-Tom’ were modified during clinorotation treatment. The ‘Micro-Tom’ plants grown under simulated microgravity exhibited a spreading growth and an increasing of the internode length. Total fruit yield, small fruit yield, leaf area, leaf dry weight, fruit dry weight, total dry weight and shoot – root ratio were lower in the clinorotated tomato plants than those grown in the control treatment. Foliar amount of carotenoids, and chlorophyll a and b were also substantially reduced under simulated microgravity conditions. Quality parameters (total soluble solids and fruit dry matter) of tomato plants were also negatively affected by clinorotation. The number of flowers per plant was increased by 32% in clinorotated plants versus controls. Fruit setting was reduced by 46% under clinorotation, while no significant difference was recorded for the pollen fertility and the seed number in small and large fruits. Clinorotation-exposed and control seeds were used in a germination trial in order to evaluate whether the seeds so formed were viable and if subsequent generations might be obtained in microgravity. Seeds formed under simulated microgravity proved to be biologically and functionally complete (germination = 78.6%) showing that ‘Micro-Tom’ plants could realize complete ontogenesis, from seed to seed in microgravity.     

The role of the tolerance–fecundity trade‐off in maintaining intraspecific seed trait variation in a widespread dimorphic herb

Coexistence of species with different seed sizes is a long‐standing issue in community ecology, and a trade‐off between fecundity and stress tolerance has been proposed to explain co‐occurrence in heterogeneous environments. Here we tested an intraspecific extension of this model: whether such trade‐off also explains seed trait variation among populations of widespread plants under stress gradients. We collected seeds from 14 populations of Plantago coronopus along the Atlantic coast in North Africa and Europe. This herb presents seed dimorphism, producing large basal seeds with a mucilaginous coat that facilitates water absorption (more stress tolerant), and small apical seeds without coats (less stress tolerant). We analysed variation among populations in number, size and mucilage production of basal and apical seeds, and searched for relationships between local environment and plant size. Populations under higher stress (higher temperature, lower precipitation, lower soil organic matter) had fewer seeds per fruit, higher predominance of basal relative to apical seeds, and larger basal seeds with thicker mucilaginous coats. These results strongly suggest a trade‐off between tolerance and fecundity at the fruit level underpins variation in seed traits among P. coronopus populations. However, seed production per plant showed the opposite pattern to seed production per fruit, and seemed related to plant size and other life‐cycle components, as an additional strategy to cope with environmental variation across the range. The tolerance–fecundity model may constitute, under stress gradients, a broader ecological framework to explain trait variation than the classical seed size–number compromise, although several fecundity levels and traits should be considered to understand the diverse strategies of widespread plants to maximise fitness in each set of local conditions.     

Cassettes for seed-specific expression tested in transformed embryogenic cultures of soybean

Soybean (Glycine max [L.] Merrill) lectin is a seed protein that accumulates in protein bodies of cotyledons during seed development. We have constructed two expression cassettes containing the 5′ and 3′ regions of the soybean lectin gene connected by aNot I restriction site. One vector also contains the 32 amino acid signal sequence. Using polymerase chain reaction (PCR), the coding region of the β-glucuronidase (uidA) gene was inserted into theNot I site of each vector. We tested the function of the expression cassettes in transformed embryogenic cultures of soybean. Development-specific GUS expression was observed in developing somatic embryos transformed with the chimeric lectin promoter-GUS constructs as determined by histochemical assays. Our data indicate that these cassettes could be used to drive expression of foreign genes to modify embryo-specific traits of soybean as protein quality or quantity in the seed.     

Seed preferences by rodents in the agri‐environment and implications for biological weed control

Post‐dispersal seed predation and endozoochorous seed dispersal are two antagonistic processes in relation to plant recruitment, but rely on similar preconditions such as feeding behavior of seed consumers and seed traits. In agricultural landscapes, rodents are considered important seed predators, thereby potentially providing regulating ecosystem services in terms of biological weed control. However, their potential to disperse seeds endozoochorously is largely unknown. We exposed seeds of arable plant species with different seed traits (seed weight, nutrient content) and different Red List status in an experimental rye field and assessed seed removal by rodents. In a complementary laboratory experiment, consumption rates, feeding preferences, and potential endozoochory by two vole species (Microtus arvalis and Myodes glareolus) were tested. Seed consumption by rodents after 24 h was 35% in the field and 90% in the laboratory. Both vole species preferred nutrient‐rich over nutrient‐poor seeds and M. glareolus further preferred light over heavy seeds and seeds of common over those of endangered plants. Endozoochory by voles could be neglected for all tested plant species as no seeds germinated, and only few intact seeds could be retrieved from feces. Synthesis and applications. Our results suggest that voles can provide regulating services in agricultural landscapes by depleting the seed shadow of weeds, rather than facilitating plant recruitment by endozoochory. In the laboratory, endangered arable plants were less preferred by voles than noxious weeds, and thus, our results provide implications for seed choice in restoration approaches. However, other factors such as seed and predator densities need to be taken into account to reliably predict the impact of rodents on the seed fate of arable plants.     

Seed storage at elevated partial pressure of oxygen, a fast method for analysing seed ageing under dry conditions

Background and Aims

Despite differences in physiology between dry and relative moist seeds, seed ageing tests most often use a temperature and seed moisture level that are higher than during dry storage used in commercial practice and gene banks. This study aimed to test whether seed ageing under dry conditions can be accelerated by storing under high-pressure oxygen.

Methods

Dry barley (Hordeum vulgare), cabbage (Brassica oleracea), lettuce (Lactuca sativa) and soybean (Glycine max) seeds were stored between 2 and 7 weeks in steel tanks under 18 MPa partial pressure of oxygen. Storage under high-pressure nitrogen gas or under ambient air pressure served as controls. The method was compared with storage at 45 °C after equilibration at 85 % relative humidity and long-term storage at the laboratory bench. Germination behaviour, seedling morphology and tocopherol levels were assessed.

Key Results

The ageing of the dry seeds was indeed accelerated by storing under high-pressure oxygen. The morphological ageing symptoms of the stored seeds resembled those observed after ageing under long-term dry storage conditions. Barley appeared more tolerant of this storage treatment compared with lettuce and soybean. Less-mature harvested cabbage seeds were more sensitive, as was the case for primed compared with non-primed lettuce seeds. Under high-pressure oxygen storage the tocopherol levels of dry seeds decreased, in a linear way with the decline in seed germination, but remained unchanged in seeds deteriorated during storage at 45 °C after equilibration at 85 % RH.

Conclusions

Seed storage under high-pressure oxygen offers a novel and relatively fast method to study the physiology and biochemistry of seed ageing at different seed moisture levels and temperatures, including those that are representative of the dry storage conditions as used in gene banks and commercial practice.     

Manipulation of saponin biosynthesis by RNA interference-mediated silencing of ��-amyrin synthase gene expression in soybean

Soybean seeds contain substantial amount of diverse triterpenoid saponins that influence the seed quality, although little is known about the physiologic functions of saponins in plants. We now describe the modification of saponin biosynthesis by RNA interference (RNAi)-mediated gene silencing targeted to β-amyrin synthase, a key enzyme in the synthesis of a common aglycon of soybean saponins. We identified two putative β-amyrin synthase genes in soybean that manifested distinct expression patterns with regard to developmental stage and tissue specificity. Given that one of these genes, GmBAS1, was expressed at a much higher level than the other (GmBAS2) in various tissues including the developing seeds, we constructed two RNAi vectors that encode self-complementary hairpin RNAs corresponding to the distinct regions of GmBAS1 under the control of a seed-specific promoter derived from the soybean gene for the α′ subunit of the seed storage protein β-conglycinin. These vectors were introduced independently into soybean. Six independent transgenic lines exhibited a stable reduction in seed saponin content, with the extent of saponin deficiency correlating with the β-amyrin synthase mRNA depletion. Although some transgenic lines produced seeds almost devoid of saponins, no abnormality in their growth was apparent and the antioxidant activity of their seeds was similar to that of control seeds. These results suggest that saponins are not required for seed development and survival, and that soybean seeds may therefore be amenable to the modification of triterpenoid saponin content and composition through molecular biologic approaches.     

Overexpression of a NTR1 in transgenic soybean confers tolerance to water stress

Methyl jasmonate (MeJA) is an important plant regulator that involves in plant development and regulates the expression of plant defense genes in response to various stresses such as wounding, drought, and pathogens. In order to determine the physiological role of endogenous MeJA in plants, a NTR1 from Brassica campestris encoding a jasmonic acid carboxyl methyltransferase that produces methyl jasmonate was constructed under the control of CaMV 35S promoter and transformed into soybean [Glycine max (L) Merrill]. The transgenic soybean plants constitutively expressed the NTR1 and accumulated more MeJA levels than wild type plants. Overexpression of the gene in transgenic soybean conferred tolerance to dehydration during seed germination and seedling growth as reflected by the percentage of the fresh weight of seedlings. In addition, the transgenic soybean plants also conferred better capacity to retain water than wild type plants when drought tolerance was tested using detached leaves.     

Natural variation in GmGBP1 promoter affects photoperiod control of flowering time and maturity in soybean

The present study screened for polymorphisms in coding and non‐coding regions of the GmGBP1 gene in 278 soybean accessions with variable maturity and growth habit characteristics under natural field conditions in three different latitudes in China. The results showed that the promoter region was highly diversified compared with the coding sequence of GmGBP1. Five polymorphisms and four haplotypes were closely related to soybean flowering time and maturity through association and linkage disequilibrium analyses. Varieties with the polymorphisms SNP_‐796G, SNP_‐770G, SNP_‐307T, InDel_‐242normal, SNP_353A, or haplotypes Hap‐3 and Hap‐4 showed earlier flowering time and maturity in different environments. The shorter growth period might be largely due to higher GmGBP1 expression levels in soybean that were caused by the TCT‐motif with SNP_‐796G in the promoter. In contrast, the lower expression level of GmGBP1 in soybean caused by RNAi interference of GmGBP1 resulted in a longer growth period under different day lengths. Furthermore, the gene interference of GmGBP1 also caused a reduction in photoperiod response sensitivity (PRS) before flowering in soybean. RNA‐seq analysis on GmGBP1 underexpression in soybean showed that 94 and 30 predicted genes were significantly upregulated and downregulated, respectively. Of these, the diurnal photoperiod‐specific expression pattern of three significant flowering time genes GmFT2a, GmFT5a, and GmFULc also showed constantly lower mRNA levels in GmGBP1‐i soybean than in wild type, especially under short day conditions. Together, the results showed that GmGBP1 functioned as a positive regulator upstream of GmFT2a and GmFT5a to activate the expression of GmFULc to promote flowering on short days.     

Soybean seed damage by different species of stink bugs

Abstract

• 1 Damage caused by the three main species of stink bugs occurring on soybean Nezara viridula (Linnaeus), Piezodorus guildinii (Westwood) and Euschistus heros (Fabricius) was compared in field cages and in greenhouses. Infestation levels of 4 stink bugs/m row of plants (field cages) and 2 stink bug/plant (greenhouse) for 15 days during the pod filling stage are reported. At harvest, the yield and seed quality were evaluated.
• 2 In the field, there was no difference in yield between infested and insect‐free plants, but damage to seed quality varied with stink bug species. Plants damaged by P. guildinii had the lowest quality seeds. From 50 g seed samples harvested in the field, the mean weight of seeds classified as ‘good’ was 37.3 g in plants infested with P. guildinii, compared to 41.8, 44.2 and 46.6 g in plants infested with E. heros, N. viridula and the control, respectively.
• 3 Plants infested with P. guildinii showed the highest number of seeds damaged by stink bugs, whereas those infested with E. heros showed the lowest damage.
• 4 Plants infested with P. guildinii had 18.5% damaged seeds, higher than the 3.6% and 3.4% damaged seeds from plants infested with the two other species and 0.1% in control plants. The percentage of non‐viable seeds due to stink bug damage was 5.7% for P. guildinii but lower for the other two species.

     

Negative correlation between altitudes and oxygen isotope ratios of seeds: exploring its applicability to assess vertical seed dispersal

Vertical seed dispersal, which plays a key role in plant escape and/or expansion under climate change, was recently evaluated for the first time using negative correlation between altitudes and oxygen isotope ratio of seeds. Although this method is innovative, its applicability to other plants is unknown. To explore the applicability of the method, we regressed altitudes on δ¹⁸O of seeds of five woody species constituting three families in temperate forests in central Japan. Because climatic factors, including temperature and precipitation that influence δ¹⁸O of plant materials, demonstrate intensive seasonal fluctuation in the temperate zone, we also evaluated the effect of fruiting season of each species on δ¹⁸O of seeds using generalized linear mixed models (GLMM). Negative correlation between altitudes and δ¹⁸O of seeds was found in four of five species tested. The slope of regression lines tended to be lower in late‐fruiting species. The GLMM analysis revealed that altitudes and date of fruiting peak negatively affected δ¹⁸O of seeds. These results indicate that the estimation of vertical seed dispersal using δ¹⁸O of seeds can be applicable for various species, not just confined to specific taxa, by identifying the altitudes of plants that produced seeds. The results also suggest that the regression line between altitudes and δ¹⁸O of seeds is rather species specific and that vertical seed dispersal in late‐fruiting species is estimated at a low resolution due to their small regression slopes. A future study on the identification of environmental factors and plant traits that cause a difference in δ¹⁸O of seeds, combined with an improvement of analysis, will lead to effective evaluation of vertical seed dispersal in various species and thereby promote our understanding about the mechanism and ecological functions of vertical seed dispersal.