Role of relative humidity, temperature, and water status in dormancy alleviation of sunflower seeds during dry after-ripening

The effect of various combinations of temperature and relative humidity on dormancy alleviation of sunflower seeds during dry after-ripening was investigated. The rate of dormancy alleviation depended on both temperature and embryo moisture content (MC). Below an embryo MC of 0.1 g H(2)O g(-1) dw, dormancy release was faster at 15 °C than at higher temperatures. This suggests that dormancy release at low MC was associated with negative activation energy, supported by Arrhenius plots, and low Q(10) values. At higher MC, the rate of dormancy alleviation increased with temperature, correlating well with the temperature dependence of biochemical processes. These findings suggests the involvement of two distinct cellular mechanisms in dormancy release; non-enzymatic below 0.1 g H(2)O g(-1) dw and associated with active metabolism above this value. The effects of temperature on seed dormancy release above the threshold MC were analysed using a population-based thermal time approach and a model predicting the rate of dormancy alleviation is provided. Sunflower embryo dormancy release was effective at temperatures above 8 °C (the base temperature for after-ripening, Tb(AR), was 8.17 °C), and the higher the after-ripening temperature above this threshold value, the higher was the rate of dormancy loss. Thermodynamic analyses of water sorption isotherms revealed that dormancy release was associated with less bound water and increased molecular mobility within the embryonic axes but not the cotyledons. It is proposed that the changes in water binding properties result from oxidative processes and can, in turn, allow metabolic activities.     

ROS production and protein oxidation as a novel mechanism for seed dormancy alleviation

At harvest, sunflower (Helianthus annuus L.) seeds are dormant and unable to germinate at temperatures below 15 degrees C. Seed storage in the dry state, known as after-ripening, is associated with an alleviation of embryonic dormancy allowing subsequent germination at suboptimal temperatures. To identify the process by which dormancy is broken during after-ripening, we focused on the role of reactive oxygen species (ROS) in this phenomenon. After-ripening entailed a progressive accumulation of ROS, namely superoxide anions and hydrogen peroxide, in cells of embryonic axes. This accumulation, which was investigated at the cellular level by electron microscopy, occurred concomitantly with lipid peroxidation and oxidation (carbonylation) of specific embryo proteins. Incubation of dormant seeds for 3 h in the presence of hydrogen cyanide (a compound that breaks dormancy) or methylviologen (a ROS-generating compound) also released dormancy and caused the oxidation of a specific set of embryo proteins. From these observations, we propose a novel mechanism for seed dormancy alleviation. This mechanism involves ROS production and targeted changes in protein carbonylation patterns.     

Variation in the loss of seed dormancy during after-ripening of wild and cultivated rice species

BACKGROUND AND AIMS: The aim of this paper was to verify the variation in the loss of seed dormancy during after-ripening and the interspecific and interpopulation variability in the degree of dormancy of seven wild and two cultivated rice species comprising 21 populations and two cultivars. METHODS: Four wild rice species from South America, Oryza glumaepatula, O. latifolia, O. grandiglumis and O. alta, and two O. sativa cultivars were tested in one experiment. In a second experiment, five wild species, O. punctata, O. eichingeri, O.rufipogon, O. latifolia and O. glumaepatula, and one cultivated species (O. glaberrima) were evaluated. Initial germination tests were performed soon after the seeds were harvested and subsequently at 2-month intervals, for a total of six storage periods in the first experiment and three in the second. All tests were conducted in the dark at a temperature of 27 degrees C. KEY RESULTS: Different patterns of after-ripening among populations within and between species were observed. CONCLUSIONS: The cultivated species (O. sativa and O. glaberrima) and, amongst the wild species, the tetraploids O. latifolia, O. grandiglumis and the diploids O. eichingeri and O. punctata, had weak dormancy, losing it completely 2 months after harvest, while O. rufipogon and O. glumaepatula exhibited pronounced dormancy. The latter showed different patterns of after-ripening between populations indigenous to the Amazon region and those originating in the Paraguay River system. Seeds of Solimoes (Amazon) and Japura origin showed weak dormancy whereas those of Paraguay origin showed deep dormancy. Ecological differences among natural habitats may be involved in such differentiation.     

Release of dormancy during after-ripening of Agrostemma githago seeds

Freshly harvested seeds of Agrostemma githago L. do not germinate when they are imbibed at 20°C. The block is located in the embryo and is relased by dry storage at 20°C (after-ripening). Freshly harvested seeds complete only a small part of the processes that occur in after-ripened seeds during the lag phase prior to germination (radicle protrusion). After-ripening removed the block on lag phase processes much faster than the block on germination. This was shown both by direct determinations of the completion of lag phase processes and by measurements of the rate of axial protein synthesis, which approximately doubles when seeds are progressing through the lag phase. It is concluded that the percentage germination does not adequately reflect the extent to which the dormancy mechanism has been overcome.     

Development of seed coat-imposed dormancy during seed maturation in Cynoglossum officinale

The relationship between seed phenolics and appearance of seed coat–imposed dormancy during seed development in Cynoglossum officinale L. was studied. Up to 24 days after anthesis, seeds failed to germinate upon imbibition in Petri dishes at 25°C. At 44 days after anthesis, seeds were fully germinable; removal of seed coats did not improve their germination or O₂ uptake. At 72 days after anthesis, mature seeds at the base of the cyme did not germinate unless their coats were removed. Removal of seed coat also stimulated O₂ uptake at this harvest date. The methanol-soluble phenolic content of the seeds increased during the early stages of seed development, in both the seed coat and the embryo. As seed development continued, the methanol-soluble phenolic content of the embryo stabilized, but that of the seed coat declined. This decline was associated with an increase in the thioglycolic acid–soluble phenolics, presumably lignins, in the seed coat. These results suggest that polymerization of methanol–soluble phenolics into lignins in the seed coat during later stages of seed development renders the seed coat of C. officinale impermeable to 03, and thus keeps the seed dormant.     

Post-harvest age-induced seed dormancy of Acer ginnala and its alleviation by growth regulator and low temperature treatments

One-month-old fruits of Acer ginnala with winged pericarp attached gave 44% germination and this was not increased by cold treatment at 4°C for 0, 10, 20, or 30 days, gibberellic acid treatment at 0, 1, 10, 100 or 1000 mg litre^-1, or ethephon treatment at 0, 2, 20, 200 or 2000 mg litre^-1. After 6 months of storage at 20–25 °C, germination of untreated fruits fell to 5% but could be restored to that of 1-month-old fruits by incubation at 4 °C for 30 days. After 9 months storage, no germination occurred in untreated fruits. Cold treatment (30 days at 4 °C partially restored germination (26%). Treatment with either gibberellic acid (1000 mg litre^-1) and 30 days at 4 °C (40%) or ethephon (100 mg litre^-] and 30 days at 4 °C improved germination (69%). The combination of all three treatments, i.e. 100 mg litre^-1 gibberellic acid, 100 mg litre^-1 ethephon and 30 days at 4 °C, optimised germination (86%). Thus, dormancy of A. ginnala developed during storage but could be reversed by a combination of treatment with low temperature and growth regulators. The highest germination (86%) was achieved after low temperature and growth regulator treatment of stored fruit.     

The self-incompatibility locus (S) and quantitative trait loci for self-pollination and seed dormancy in sunflower

Wild populations of common sunflower (Helianthus annuus L.) are self-incompatible and have deep seed dormancy, whereas modern cultivars, inbreds, and hybrids are self-compatible and partially-to-strongly self-pollinated, and have shallow seed dormancy. Self-pollination (SP) and seed dormancy are genetically complex traits, the number of self-compatibility (S) loci has been disputed, and none of the putative S loci have been genetically mapped in sunflower. We genetically mapped quantitative trait loci (QTL) for self-incompatibility (SI), SP, and seed dormancy in a backcross population produced from a cross between an elite, self-pollinated, nondormant inbred line (NMS373) and a wild, self-incompatible, dormant population (ANN1811). A population consisting of 212 BC₁ progeny was subsequently produced by backcrossing a single hybrid individual to NMS373. BC₁ progeny produced 0–838 seeds per primary capitula when naturally selfed and 0–518 seeds per secondary capitula when manually selfed and segregated for a single S locus. The S locus mapped to linkage group 17 and was tightly linked to a cluster of previously identified QTL for several domestication and postdomestication traits. Two synergistically interacting QTL were identified for SP among self-compatible (ss) BC₁ progeny (R²=34.6%). NMS373 homozygotes produced 271.5 more seeds per secondary capitulum than heterozygotes. Germination percentages of seeds after-ripened for 4 weeks ranged from 0% to 100% among self-compatible BC₁S₁ families. Three QTL for seed dormancy were identified (R²=38.3%). QTL effects were in the predicted direction (wild alleles decreased self-pollination and seed germination). The present analysis differentiated between loci governing SI and SP and identified DNA markers for bypassing SI and seed dormancy in elite × wild crosses through marker-assisted selection.Electronic Supplementary Material Electronic supplementary material is available for this article at     

Cell lineage patterns in the shoot meristem of the sunflower embryo in the dry seed

We mapped the fate of cells in the shoot meristem of the dry-seed embryo of sunflower, Helianthus annuus L. cv. Peredovic, using irradiation-induced somatic sectors. We analyzed 249 chlorophyll-deficient or glabrous (hairless) sectors generated in 236 plants. Most sectors observed in the inflorescence extended into vegetative nodes. Thus cell lineages that ultimately gave rise to reproductive structures also contributed to vegetative structures. No single sector extended the entire length of the shoot. Thus the shoot is not derived from one or a few apical initials. Rather, the position, vertical extent, and width of the sectors at different levels of the shoot suggest that the shoot is derived from three to four circumferential populations of cells in each of three cell layers of the embryo meristem. Sectors had no common boundaries even in plants with two or three independent sectors, but varied in extent and overlapped along the length of the shoot. Thus individual cells in a single circumferential population behaved independently to contribute lineages of different vertical extents to the growing shoot. The predicted number of circumferential populations of cells as well as the apparent cell number in each population was consistent with the actual number of cells in the embryo meristem observed in histological sections.     

Polypeptide markers for low temperature stress during seed germination in sunflower

Sunflower seeds behaved as chilling and freezing sensitive and also exhibited acclimation under low seed moisture content (< 1 %). At high seed moisture content (approx. 22 %) they tolerated chilling stress but failed to acclimate under freezing temperatures. Pre-imbibitional chilling (5 °C) or freezing (−5 or −10 °C) stress significantly enhanced total soluble protein (TSP) content. Chilling treatment after imbibition (in contrast to pre-imbibition) enhanced germination and this was accompanied by increase in 30, 24 and 21.9 kDa TSPs content (3 d after germination). Freezing at −5 and −10 °C suppressed seed germination and increased content of 78 and 56.2 kDa wall bound proteins. Chilling acclimation decreased 35.4, 33.9, 29.5, 23.4 and 21.4 kDa TSPs.     

ZBP1 regulates mRNA stability during cellular stress

An essential constituent of the integrated stress response (ISR) is a reversible translational suppression. This mRNA silencing occurs in distinct cytoplasmic foci called stress granules (SGs), which transiently associate with processing bodies (PBs), typically serving as mRNA decay centers. How mRNAs are protected from degradation in these structures remains elusive. We identify that Zipcode-binding protein 1 (ZBP1) regulates the cytoplasmic fate of specific mRNAs in nonstressed cells and is a key regulator of mRNA turnover during the ISR. ZBP1 association with target mRNAs in SGs was not essential for mRNA targeting to SGs. However, ZBP1 knockdown induced a selective destabilization of target mRNAs during the ISR, whereas forced expression increased mRNA stability. Our results indicate that although targeting of mRNAs to SGs is nonspecific, the stabilization of mRNAs during cellular stress requires specific protein-mRNA interactions. These retain mRNAs in SGs and prevent premature decay in PBs. Hence, mRNA-binding proteins are essential for translational adaptation during cellular stress by modulating mRNA turnover.     

Processing of N-glycans of two yellow lupin phosphohydrolases during seed maturation and dormancy

Acid phosphatase (AP) and diphosphonucleoside phosphatase/phosphodiesterase (PPD1) were purified from yellow lupin (Lupinus luteus L.) immature green seeds (40 days after blooming), dry seeds (40 days later) and dry seeds stored for 160 days. Both enzymes are known to differ in the type of N-glycosylation: the first has an N-glycosylation pattern typical for a vacuolar protein, while the second enzyme has a pattern typical for an extracellular or membrane-bound protein. N-Glycans were released from each of the enzyme preparations, fluorescence labeled, separated and identified by HPLC (GlycoSep N and GlycoSep H columns). Changes in the level of each N-glycan during seed maturation and dormancy were compared. The results show that N-glycan processing in the case of AP and PPD1-two proteins residing in the same plant organ, but possibly in different compartments-is not synchronized and performed not only in metabolically active maturing seeds, but also in metabolically inactive dormant seeds.     

Endo-β-mannanase activity during dormancy alleviation and germination of white spruce (Picea glauca) seeds

It is not known how embryos of seeds of the Pinaceae protrude from their enclosing tissues to complete germination. Prior to protrusion of the radicle there is an increase in endo-β-1,4-mannanase (EC 3.2.1.78) activity associated with weakening of the micropylar megagametophyte/nucellus from seeds of white spruce ( Picea glauca [Moench.] Voss). Mannanase activity is present as three isoforms (pI values 5.0, 4.8, 4.7) in both the embryo and surrounding structures (megagametophyte and nucellus) prior to and during imbibition. Activity of all the isoforms increases in the chalazal and micropylar megagametophyte during germination. Activity then declines after the testa splits, typically 1 day prior to radicle protrusion, due partially to its leaching from the seed into the surrounding water. Activity increases in the cotyledons and axis as the embryo commences elongation. Seeds from dormant seedlots exhibit a lower germination percentage, relative to seeds from nondormant seedlots, and the force necessary for the embryo to puncture the surrounding structures tends to be greater. Although similar mannanase activities are present in unimbibed seeds of dormant and nondormant seedlots, during germination, enzyme activity in seeds of dormant seedlots is lower. Moist chilling alleviates dormancy in the seeds of the Pinaceae and, during 3 weeks of this treatment, mannanase activity slowly increases. After 3 weeks of moist chilling and regardless of whether the seedlot was dormant or not prior to moist chilling, the force necessary to puncture the micropylar megagametophyte and nucellus is lower, and the speed of germination greater. Seeds from previously dormant seedlots also complete germination to a greater percentage, relative to unchilled seeds from dormant seedlots. Upon transfer to 25°C, mannanase activity in moist-chilled seeds decreases during germination of all seedlots regardless of their previous dormancy status.