Studies in Wild Oat Seed Dormancy: I. THE ROLE OF ETHYLENE IN DORMANCY BREAKAGE AND GERMINATION OF WILD OAT SEEDS (AVENA FATUA L.)

Seed of Avena fatua were shown to exhibit a characteristic loss of dormancy during dry storage at 25 C, whereas similar seed stored at 5 C maintained dormancy. 2-Chloroethylphosphonic acid was shown to increase germination of partly dormant seed imbibed under certain temperature regimes; a similar effect could not be established for fully dormant or fully nondormant seed. Using gas-liquid chromatography, natural ethylene levels were followed during imbibition of fully dormant and nondormant seed. A large peak in production was observed in the period prior to radicle emergence in the case of the nondormant seed. Measurements of ethylene production taken at 15 C, following periods of after-ripening in moist soil at either 5 or 25 C, indicated that endogenous production was unlikely to be a main cause of dormancy breakage in this species. The possibility that endogenous ethylene could play a role in natural dormancy breakage in aged seeds is discussed. The practical possibilities of 2-chloroethylphosphonic acid as a dormancy breaking agent in a field situation are outlined.     

Tanned or Burned: The Role of Fire in Shaping Physical Seed Dormancy

Plant species with physical seed dormancy are common in mediterranean fire-prone ecosystems. Because fire breaks seed dormancy and enhances the recruitment of many species, this trait might be considered adaptive in fire-prone environments. However, to what extent the temperature thresholds that break physical seed dormancy have been shaped by fire (i.e., for post-fire recruitment) or by summer temperatures in the bare soil (i.e., for recruitment in fire-independent gaps) remains unknown. Our hypothesis is that the temperature thresholds that break physical seed dormancy have been shaped by fire and thus we predict higher dormancy lost in response to fire than in response to summer temperatures. We tested this hypothesis in six woody species with physical seed dormancy occurring in fire-prone areas across the Mediterranean Basin. Seeds from different populations of each species were subject to heat treatments simulating fire (i.e., a single high temperature peak of 100°C, 120°C or 150°C for 5 minutes) and heat treatments simulating summer (i.e., temperature fluctuations; 30 daily cycles of 3 hours at 31°C, 4 hours at 43°C, 3 hours at 33°C and 14 hours at 18°C). Fire treatments broke dormancy and stimulated germination in all populations of all species. In contrast, summer treatments had no effect over the seed dormancy for most species and only enhanced the germination in Ulex parviflorus, although less than the fire treatments. Our results suggest that in Mediterranean species with physical dormancy, the temperature thresholds necessary to trigger seed germination are better explained as a response to fire than as a response to summer temperatures. The high level of dormancy release by the heat produced by fire might enforce most recruitment to be capitalized into a single post-fire pulse when the most favorable conditions occur. This supports the important role of fire in shaping seed traits.     

Environmental regulation of dormancy loss in seeds of Lomatium dissectum (Apiaceae)

Background and Aims

Lomatium dissectum (Apiaceae) is a perennial, herbaceous plant of wide distribution in Western North America. At the time of dispersal, L. dissectum seeds are dormant and have under-developed embryos. The aims of this work were to determine the requirements for dormancy break and germination, to characterize the type of seed dormancy, and to determine the effect of dehydration after embryo growth on seed viability and secondary dormancy.

Methods

The temperature requirements for embryo growth and germination were investigated under growth chamber and field conditions. The effect of GA₃ on embryo growth was also analysed to determine the specific type of seed dormancy. The effect of dehydration on seed viability and induction of secondary dormancy were tested in seeds where embryos had elongated about 4-fold their initial length. Most experiments examining the nature of seed dormancy were conducted with seeds collected at one site in two different years. To characterize the degree of variation in dormancy-breaking requirements among seed populations, the stratification requirements of seeds collected at eight different sites were compared.

Key Results

Embryo growth prior to and during germination occurred at temperatures between 3 and 6 °C and was negligible at stratification temperatures of 0·5 and 9·1 °C. Seeds buried in the field and exposed to natural winter conditions showed similar trends. Interruption of the cold stratification period by 8 weeks of dehydration decreased seed viability by about 30 % and induced secondary dormancy in the remaining viable seeds. Comparison of the cold stratification requirements of different seed populations indicates that seeds collected from moist habitats have longer cold stratification requirements that those from semiarid environments.

Conclusions

Seeds of L. dissectum have deep complex morphophysiological dormancy. The requirements for dormancy break and germination reflect an adaptation to trigger germination in late winter.Key words: Apiaceae, cold stratification, Lomatium dissectum, morphophysiological dormancy, secondary dormancy, seed germination     

Hormonal and temperature regulation of seed dormancy and germination in <Emphasis Type="Italic">Leymus chinensis</Emphasis>

Fluctuating temperature plays a critical role in determining the timing of seed germination in many plant species. However, the physiological and biochemical mechanisms underlying such a response have been paid little attention. The present study investigated the effect of plant growth regulators and cold stratification in regulating Leymus chinensis seed germination and dormancy response to temperature. Results showed that seed germination was less than 2 % at all constant temperatures while fluctuating temperature significantly increased germination percentage. The highest germination was 71 % at 20/30 °C. Removal of the embryo enclosing material of L. chinensis seed germinated to 74 %, and replaced the requirement for fluctuating temperature to germinate, by increasing embryo growth potential. Applications of GA₄₊₇ significantly increased seed germination at constant temperature. Also, inhibition of GA biosynthesis significantly decreased seed germination at fluctuating temperatures depending upon paclobutrazol concentration. This implied GA was necessary for non-dormant seed germination and played an important role in regulating seed germination response to temperature. Inhibition of ABA biosynthesis during imbibition completely released seed dormancy at 20/30 °C, but showed no effect on seed germination at constant temperature, suggesting ABA biosynthesis was important for seed dormancy maintenance but may not involve in seed germination response to temperature. Cold stratification with water or GA₃ induced seed into secondary dormancy, but this effect was reversed by exogenous FL, suggesting ABA biosynthesis during cold stratification was involved in secondary dormancy. Also, cold stratification with FL entirely replaced the requirement of fluctuating temperature for germination with seeds having 73 % germination at constant temperature. This appears to be attributed to inhibition of ABA biosynthesis and an increase of GA biosynthesis during cold stratification, leading to an increased embryo growth potential. We suggest that fluctuating temperature promotes seed germination by increasing embryo growth potential, mainly attributed to GA biosynthesis during imbibitions. ABA is important for seed dormancy maintenance and induction but showed less effect on non-dormant seed germination response to temperature.     

Seed dormancy varies widely among Arabidopsis thaliana populations both between and within Fennoscandia and Italy

The timing of germination is a key life‐history trait in plants, which is strongly affected by the strength of seed dormancy. Continental‐wide genetic variation in seed dormancy has been related to differences in climate and the timing of conditions suitable for seedling establishment. However, for predictions of adaptive potential and consequences of climatic change, information is needed regarding the extent to which seed dormancy varies within climatic regions and the factors driving such variation.We quantified dormancy of seeds produced by 17 Italian and 28 Fennoscandian populations of Arabidopsis thaliana when grown in the greenhouse and at two field sites in Italy and Sweden. To identify possible drivers of among‐population variation in seed dormancy, we examined the relationship between seed dormancy and climate at the site of population origin, and between seed dormancy and flowering time.Seed dormancy was on average stronger in the Italian compared to the Fennoscandian populations, but also varied widely within both regions. Estimates of seed dormancy in the three maternal environments were positively correlated. Among Fennoscandian populations, seed dormancy tended to increase with increasing summer temperature and decreasing precipitation at the site of population origin. In the smaller sample of Italian populations, no significant association was detected between mean seed dormancy and climate at the site of origin. The correlation between population mean seed dormancy and flowering time was weak and not statistically significant within regions.The correlation between seed dormancy and climatic factors in Fennoscandia suggests that at least some of the among‐population variation is adaptive and that climate change will affect selection on this trait. The weak correlation between population mean seed dormancy and flowering time indicates that the two traits can evolve independently.     

Fire and summer temperatures interact to shape seed dormancy thresholds

Background and AimsIn Mediterranean ecosystems, the heat shock of wildfire disrupts physical seed dormancy in many plant species. This triggers germination in the post-fire environment where seedling establishment is optimal due to decreased competition and increased resource availability. However, to maintain the soil seed bank until a fire occurs, the minimum heat capable of breaking seed dormancy (i.e. the lower heat threshold) must be above the maximum temperatures typically observed in the soil during the summer. We therefore hypothesized that summer temperatures have shaped heat requirements for physical dormancy release. Specifically, we predicted that seeds from populations growing under warmer summers will have higher values of the lower heat threshold.MethodsTo evaluate this prediction, we collected seeds from two Cistus species in 31 populations (20 Cistus albidus and 11 Cistus salviifolius) along a climate gradient of summer temperatures on the eastern coast of Spain. For each population, seeds were treated to 10 min heat shocks, from 30 to 120 °C in 5 °C increments (19 treatments), to simulate increasing heat doses from summer to fire-related temperatures. Seeds were then germinated in the lab.Key ResultsFor all populations, maximum germination was observed when applying temperatures associated with fire. Lower heat thresholds varied among populations, with a positive relationship between summer temperatures at seed population origin and the heat dose required to break dormancy.ConclusionsOur results suggest that fire drives maximum dormancy release for successful post-fire germination, while summer temperatures determine lower heat thresholds for ensuring inter-fire seed bank persistence. Significant among-population variation of thresholds also suggests that post-fire seeder species have some potential to modify their dormancy release requirements in response to changing climate.     

Germination responses to temperature and water potential in Jatropha curcas seeds: a hydrotime model explains the difference between dormancy expression and dormancy induction at different incubation temperatures

Background and Aims

Jatropha curcas is a drought-resistant tree whose seeds are a good source of oil that can be used for producing biodiesel. A successful crop establishment depends on a rapid and uniform germination of the seed. In this work we aimed to characterize the responses of J. curcas seeds to temperature and water availability, using thermal time and hydrotime analysis,

Methods

Thermal and hydrotime analysis was performed on germination data obtained from the incubation of seeds at different temperatures and at different water potentials.

Key Results

Base and optimum temperatures were 14·4 and 30 °C, respectively. Approximately 20 % of the seed population displayed absolute dormancy and part of it displayed relative dormancy which was progressively expressed in further fractions when incubation temperatures departed from 25 °C. The thermal time model, but not the hydrotime model, failed to describe adequately final germination percentages at temperatures other than 25 °C. The hydrotime constant, θ_H, was reduced when the incubation temperature was increased up to 30 °C, the base water potential for 50 % germination,Ψ_b(50), was less negative at 20 and 30 °C than at 25 °C, indicating either expression or induction of dormancy. At 20 °C this less negative Ψ_b(50) explained satisfactorily the germination curves obtained at all water potentials, while at 30 °C it had to be corrected towards even less negative values to match observed curves at water potentials below 0. Hence, Ψ_b(50) appeared to have been further displaced to less negative values as exposure to 30 °C was prolonged by osmoticum. These results suggest expression of dormancy at 20 °C and induction of secondary dormancy above 25 °C. This was confirmed by an experiment showing that inhibition of germination imposed by temperatures higher than 30 °C, but not that imposed at 20 °C, is a permanent effect.

Conclusions

This study revealed (a) the extremely narrow thermal range within which dormancy problems (either through expression or induction of dormancy) may not be encountered; and (b) the high sensitivity displayed by these seeds to water shortage. In addition, this work is the first one in which temperature effects on dormancy expression could be discriminated from those on dormancy induction using a hydrotime analysis.     

A transgenic approach to controlling wheat seed dormancy level by using Triticeae DOG1-like genes

Seed dormancy is an important agronomic trait: low levels can cause premature germination, while too much can inhibit uniform germination. As an approach to controlling the seed dormancy level in crops, we used Triticeae DOG1-like genes as transgenes. DOG1 is an Arabidopsis gene that underlies natural variation in seed dormancy. We previously showed that although their sequence similarities to DOG1 were low, some cereal DOG1-like genes enhanced seed dormancy in Arabidopsis. Here, we introduced two DOG1-like genes, TaDOG1L4 from wheat and HvDOG1L1 from barley, individually into the wheat cultivar Fielder. Their overexpression under the control of a maize ubiquitin promoter enhanced the seed dormancy level while leaving other traits unchanged. TaDOG1L4 was more effective than HvDOG1L1, which accords with the previously revealed difference in the effectiveness of these two genes in Arabidopsis seed dormancy. Knockdown of endogenous TaDOG1L4 in Fielder using double-strand RNA interference decreased the seed dormancy level by several tens of percent. This result indicates that some degree of seed dormancy inherent in wheat is imparted by DOG1-like genes.     

Dormancy release and seed ageing in the endangered species <Emphasis Type="Italic">Silene diclinis</Emphasis>

The influence of seed testa color, temperature and seed water content on dormancy release and seed viability loss in the endangered, endemic species Silene diclinis (Lag.) M. Laínz was evaluated. Dormant heterogeneous seeds (black, red and grey colored) were exposed to three different temperatures (5, 20, and 35°C) and two relative humidities (33 and 60%) in order to assay their dormancy release. Longevity behavior was studied for the three colored seeds, storing samples at nine different combinations of temperature (5, 20 and 35°C) and relative humidities (33, 60 and 90%). According to our findings, seed heteromorphism was not related to neither break of dormancy nor seed storage behavior. Silene diclinis seeds present dormancy after collection, and need an after-ripening period to germinate. Temperature and relative humidity are positively correlated with dormancy release and seed ageing. Therefore, both factors must be carefully controlled during seed manipulation in the laboratory for long term seed conservation purposes. When seeds are stored immediately after collection (dormant), if the temperature of storage is above the base temperature for dormancy release found in this work (between 2.7 and 1.6°C), seeds may eventually overcome dormancy. On the other hand if seeds are stored after an after-ripening period, storage at low temperature does not induce secondary dormancy.     

Polymorphic Homoeolog of Key Gene of RdDM Pathway,ARGONAUTE4_9 class Is Associated with Pre-Harvest Sprouting in Wheat (Triticum aestivum L.)

Resistance to pre-harvest sprouting (PHS) is an important objective for the genetic improvement of many cereal crops, including wheat. Resistance, or susceptibility, to PHS is mainly influenced by seed dormancy, a complex trait. Reduced seed dormancy is the most important aspect of seed germination on a spike prior to harvesting, but it is influenced by various environmental factors including light, temperature and abiotic stresses. The basic genetic framework of seed dormancy depends on the antagonistic action of abscisic acid (ABA) and gibberellic acid (GA) to promote dormancy and germination. Recent studies have revealed a role for epigenetic changes, predominantly histone modifications, in controlling seed dormancy. To investigate the role of DNA methylation in seed dormancy, we explored the role of ARGONAUTE4_9 class genes in seed development and dormancy in wheat. Our results indicate that the two wheat AGO4_9 class genes i.e. AGO802 and AGO804 map to chromosomes 3S and 1S are preferentially expressed in the embryos of developing seeds. Differential expressions of AGO802-B in the embryos of PHS resistant and susceptible varieties also relates with DNA polymorphism in various wheat varieties due to an insertion of a SINE-like element into this gene. DNA methylation patterns of the embryonic tissue from six PHS resistant and susceptible varieties demonstrate a correlation with this polymorphism. These results suggest a possible role for AGO802-B in seed dormancy and PHS resistance through the modulation of DNA methylation.     

Effects of Stratification, Gibberellic acid and Germination Temperature on the Germination of Betula nana

Experiments were carried out with three seed lots of Betula nana collected in 1967 from different localities in Norway. Seeds were stratified for 0-20 days in dark at +2-+3 °C on filter papers moistened with distilled water, or treated with solution of GA₃ for 24 h at room temperature, and then moved into special germination boxes that were placed in different temperature conditions. All the seed lots had conditional dormancy. Quantitatively, the dormancy was different in the different seed lots (pronenances), but there were no qualitative difference in the reaction to stratification gibberellic acid and to germination temperature. Differences between seed lots may have been due to different stage of seed development. The dormancy was deepest at low temperatures(12 and 15°C) decreasing gradually with increasing temperature (to 24 °C). The dormancy was effectively broken by a short stratification (from 5 to 15 days), and by treatment with gibberellic acid. The deeper the dormancy and the lower the germination temperature the longer the stratification that was needed for maximum germination. Similarly, the concentration of GA₃ needed for maximum germination increased with decreasing temperature and with increasing dormancy.     

A local dormancy cline is related to the seed maturation environment,population genetic composition and climate

Background and Aims

Seed dormancy varies within species in response to climate, both in the long term (through ecotypes or clines) and in the short term (through the influence of the seed maturation environment). Disentangling both processes is crucial to understand plant adaptation to environmental changes. In this study, the local patterns of seed dormancy were investigated in a narrow endemic species, Centaurium somedanum, in order to determine the influence of the seed maturation environment, population genetic composition and climate.

Methods

Laboratory germination experiments were performed to measure dormancy in (1) seeds collected from different wild populations along a local altitudinal gradient and (2) seeds of a subsequent generation produced in a common garden. The genetic composition of the original populations was characterized using intersimple sequence repeat (ISSR) PCR and principal co-ordinate analysis (PCoA), and its correlation with the dormancy patterns of both generations was analysed. The effect of the local climate on dormancy was also modelled.

Key Results

An altitudinal dormancy cline was found in the wild populations, which was maintained by the plants grown in the common garden. However, seeds from the common garden responded better to stratification, and their release from dormancy was more intense. The patterns of dormancy variation were correlated with genetic composition, whereas lower temperature and summer precipitation at the population sites predicted higher dormancy in the seeds of both generations.

Conclusions

The dormancy cline in C. somedanum is related to a local climatic gradient and also corresponds to genetic differentiation among populations. This cline is further affected by the weather conditions during seed maturation, which influence the receptiveness to dormancy-breaking factors. These results show that dormancy is influenced by both long-and short-term climatic variation. Such processes at such a reduced spatial scale highlight the potential of plants to adapt to fast environmental changes.     

The absence of histone H2B monoubiquitination in the Arabidopsis hub1 (rdo4) mutant reveals a role for chromatin remodeling in seed dormancy

Seed dormancy is defined as the failure of a viable seed to germinate under favorable conditions. Besides playing an adaptive role in nature by optimizing germination to the most suitable time, a tight control of dormancy is important in crop plants. Extensive genetic and physiological studies have identified the involvement of several factors, but the molecular mechanisms underlying this process are still largely unknown. We cloned the HISTONE MONOUBIQUITINATION1 (HUB1) gene, of which the mutant (previously identified as reduced dormancy4) has reduced seed dormancy and several pleiotropic phenotypes. HUB1 encodes a C3HC4 RING finger protein. The Arabidopsis thaliana genome contains one HUB1 homolog, which we named HUB2. The hub2 mutant also has reduced seed dormancy and is not redundant with hub1. Homologs of HUB1 and HUB2 in other species are required for histone H2B monoubiquitination. In agreement with this, the ubiquitinated form of histone H2B could not be detected in the hub1 and hub2 mutants. In yeast and human cells, histone H2B monoubiquitination is associated with actively transcribed genes. The hub1 mutant showed altered expression levels for several dormancy-related genes. We propose a role for chromatin remodeling in seed dormancy by H2B monoubiquitination through HUB1 and HUB2.     

Ecology of seed dormancy and germination in sedges (Carex)

The genus Carex, with its wide distribution and large number of species yet with a rather uniform life history, is a very convenient group for comparative studies of germination ecology at the generic level. The combination of a strict or conditional primary dormancy, a light requirement for germination, low germination at constant temperatures, a positive response to diurnal temperature fluctuations and an induction of secondary dormancy in late spring by increasing environmental temperatures are attributes that were found to be characteristics shared by almost all the Carex species investigated, though there was variation between species in the degree to which these characters were expressed. In almost all species, dormancy was broken by stratification at low temperatures, though few species gained the ability to germinate at temperatures <10 °C. There is evidence that long-term physiological changes and the structure of seed coats can play a decisive role in delaying germination. High dormancy levels were found mainly in Carices with large seeds (>0.9 mg), probably due to a thicker seed coat and hence a higher resistance to germination. Differences in primary dormancy between sedges of various habitats could not be established. However, there was a tendency for temperature limits to be low in forest sedges. Many species of wetlands and open sites showed a greater capability to respond to fluctuating temperatures than species of dry sites. These dormancy and germination traits not only enable the accumulation of seeds in the soil, but also constitute seasonal seed regeneration strategies that rely on the high longevity of seeds and the formation of persistent seed banks. Temperate Carices are mainly adapted to exploit the temporally and spatially infrequent occurrence of canopy gaps that become available only in late spring or early summer, whereas the colonization of gaps at the beginning of the vegetation period is largely prevented by a high temperature requirement for germination. Many of the dormancy and germination characteristics of Carices are important in Cyperaceae generally. A greater diversity of germination responses, however, can be found in the related families, Juncaceae and Poaceae. Our present knowledge is not sufficient to determine unequivocally whether a phylogenetic component contributes significantly to the germination behaviour of the genus Carex, but certain tendencies are clearly indicated.     

Seed Maturation Regulators Are Related to the Control of Seed Dormancy in Wheat (Triticum aestivum L.)

In Arabidopsis, the regulation network of the seed maturation program controls the induction of seed dormancy. Wheat EST sequences showing homology with the master regulators of seed maturation, LEAFY COTYLEDON1 (LEC1), LEC2 and FUSCA3 (FUS3), were searched from databases and designated respectively as TaL1L (LEC1-LIKE), TaL2L (LEC2-LIKE), and TaFUS3. TaL1LA, TaL2LA and TaFUS3 mainly expressed in seeds or embryos, with the expression limited to the early stages of seed development. Results show that tissue-specific and developmental-stage-dependent expressions are similar to those of seed maturation regulators in Arabidopsis. In wheat cultivars, the expression level of TaL1LA is correlated significantly with the germination index (GI) of whole seeds at 40 days after pollination (DAP) (r = –0.83**). Expression levels of TaFUS3 and TaL2LA are significantly correlated respectively with GIs at 40 DAP and 50 DAP, except for dormant cultivars. No correlation was found between the expression level of TaVP1, orthologue of ABA INSENSITIVE3 (ABI3), and seed dormancy. DELAY OF GERMINATION1 (DOG1) was identified as a quantitative trait locus (QTL) for the regulation of seed dormancy in Arabidopsis. Its promoter has RY motif, which is a target sequence of LEC2. Significant correlation was found between the expression of TaDOG1 and seed dormancy except for dormant cultivars. These results indicate that TaL1LA, TaL2LA, and TaFUS3 are wheat orthologues of seed maturation regulators. The expressions of these genes affect the level of seed dormancy. Furthermore, the pathways, which involve seed maturation regulators and TaDOG1, are important for regulating seed dormancy in wheat.