Seed dormancy responses to temperature relate to Nothofagus species distribution and determine temporal patterns of germination across altitudes in Patagonia.

Seeds integrate environmental cues that modulate their dormancy and germination. Although many mechanisms have been identified in laboratory experiments, their contribution to germination dynamics in existing communities and their involvement in defining species habitats remain elusive. By coupling mathematical models with ecological data we investigated the contribution of seed temperature responses to the dynamics of germination of three Nothofagus species that are sharply distributed across different altitudes in the Patagonian Andes. Seed responsiveness to temperature of the three Nothofagus species was linked to the thermal characteristics of their preferred ecological niche. In their natural distribution range, there was overlap in the timing of germination of the species, which was restricted to mid-spring. By contrast, outside their species distribution range, germination was temporally uncoupled with altitude. This phenomenon was described mathematically by the interplay between interspecific differences in seed population thermal parameters and the range in soil thermic environments across different altitudes. The observed interspecific variations in seed responsiveness to temperature and its environmental regulation, constitute a major determinant of the dynamics of Nothofagus germination across elevations. This phenomenon likely contributes to the maintenance of patterns of species abundance across altitude by placing germinated seeds in a favorable environment for plant growth.

Molecular mechanisms underlying the entrance in secondary dormancy of Arabidopsis seeds.

As seasons change, dormant seeds cycle through dormant states until the environmental conditions are favourable for seedling establishment. Dormancy cycle is widespread in the plant kingdom allowing the seeds to display primary and secondary dormancy. Several reports in the last decade have focused on understanding the molecular mechanisms of primary dormancy, but our knowledge regarding secondary dormancy is limited. Here, we studied secondary dormancy induced in Arabidopsis thaliana by incubating seeds at 25 °C in darkness for 4 d. By physiological, pharmacological, expression and genetics approaches, we demonstrate that (1) the entrance in secondary dormancy involves changes in the content and sensitivity to GA, but the content and sensitivity to ABA do not change, albeit ABA is required; (2) RGL2 promotes the entrance in secondary dormancy through ABI5 action; and (3) multivariate analysis with 18 geographical and environmental parameters of accession collection place suggests that temperature is an important variable influencing the induction of secondary dormancy in nature.

Transcriptional programs related to phytochrome A function in Arabidopsis seed germination.

In Arabidopsis seeds, germination is promoted only by phytochromes, principally phytochrome B (phyB) and phytochrome A (phyA). Despite the abundant information concerning the molecular basis of phyB signaling downstream of PIF1/PIL5, the signaling network inducing germination by phyA is poorly known. Here, we describe the influence of phyA on the transcriptome of Arabidopsis seeds when germination is induced by a far-red (FR) pulse. The expression of 11% of the genome was significantly regulated by phyA. Most of the genes were up-regulated and the changes noted late (i.e. 5 h after a FR pulse), whereas changes in down-regulated genes were more abundant earlier (i.e. 0.5 h after a FR pulse). Auxin- and GA-associated elements were overrepresented in the genes that were modified by phyA. A significant number of genes whose expression was affected by phyA had not been previously reported to be dependent on PIL5. Among them, homozygotic mutant seeds of MYB66, a SAUR-like protein, PIN7, and GASA4 showed an impaired promotion of germination by phyA. Natural variation at the transcriptional level was found in early signaling and GA metabolic genes, but not in ABA metabolic and expansin genes between Columbia and Landsberg erecta accessions. Although phyA and phyB/PIL5 signaling pathways share some molecular components, our data suggest that phyA signaling is partially independent of PIL5 when germination is promoted by very low fluences of light.

Gregarious bamboo flowering opens a window of opportunity for regeneration in a temperate forest of Patagonia.

Rare gregarious flowering of understorey bamboo species occurs in temperate and subtropical forests around the world, but the ecological consequences of this phenomenon for forest regeneration are not well understood.Field experiments were conducted in an old-growth temperate forest in Patagonia,Argentina after a massive bamboo flowering event, to examine whether light quality and other changes in microhabitats could affect seed germination and growth of overstorey species. Germination of southern beech (Nothofagus obliqua) was positively correlated with red:far red (R:FR) ratios in a range of microhabitats generated by the death of the understorey bamboo (Chusquea culeou). Experimental modification of understorey R:FR ratios to mimic alternative light environments reversed this germination response in plots with senescent understorey, but not in plots with live bamboo. Laboratory incubations demonstrated a significant interaction between R:FR ratios and thermal amplitude in promoting seed germination. Microhabitats also significantly affected the growth of emerged seedlings. Microenvironmental changes generated by this flowering event appear to have opened a window of opportunity for germination and growth of overstorey species.We demonstrate that natural gradients in light quality associated with this ecological phenomenon are a major component affecting forest regeneration in this ecosystem.

Light-related loci controlling seed germination in Ler x Cvi and Bay-0 x Sha recombinant inbred-line populations of Arabidopsis thaliana.

BACKGROUND AND AIMS: Dormancy is a complex trait finely regulated by hormones and environmental factors. The phytochromes that sense red:far-red (R:FR) are the sole photoreceptors involved in the termination of dormancy and the induction of germination by light. The aims of this study were to identify and characterize loci controlling this process in seeds of Arabidopsis thaliana. METHODS: Recombinant inbred lines (RILs) derived from Landsberg erecta and Cape Verde Islands (Ler x Cvi), and Bayreuth and Shahdara (Bay-0 x Sha) were used to map loci related to light effects in seeds previously exposed to chilling and after-ripening periods. KEY RESULTS: Substantial genetic variation was found between accessions of A. thaliana in the induction of germination by light. Twelve loci were identified under R, FR or darkness, some of which were novel loci: DOG8, DOG9, DOG13, DOG14 and DOG15 detected in the Ler x Cvi RIL population; and DOG10, DOG11 and DOG12 mapped in the Bay-0 x Sha RIL population. Furthermore, independent loci were mapped for the induction of germination by low fluence (DOG-LF1 and DOG-LF2) and very low fluence of light (DOG-VLF1) in the Ler x Cvi RIL population. Several loci were confirmed and characterized after different after-ripening and chilling treatments through near-isogenic lines (NILs) and heterogeneous inbred families (HIFs). CONCLUSIONS: The results show that one group of loci act in a wide range of environmental scenarios, whereas a smaller group of loci are relevant only under a narrower set of conditions when the influence of the most-prevalent loci is reduced as a consequence of changes in the physiological status of the seeds. In addition, the identification of specific loci controlling the action modes of the phytochromes improves our understanding of the two independent signalling pathways that promote germination in response to light.

The very-low-fluence and high-irradiance responses of the phytochromes have antagonistic effects on germination, mannan-degrading activities, and DfGA3ox transcript levels in Datura ferox seeds.

Seed germination can be promoted by the modes of action of two of the phytochromes: the low-fluence response (LFR), which is the classical red (R)-far-red (FR) reversible response and the very-low-fluence response (VLFR) that can be saturated by extremely low levels of Pfr, which can be elicited by a saturating FR pulse. The Datura ferox seed population used in this work had acquired the capacity to germinate through a VLFR after pretreatment in a water-saturated atmosphere (WSA) at constant 25 degrees C. After 12 d in WSA germination after a FR pulse was 82%, while it was less than 10% in darkness. It was found that the VLFR of germination is associated with increments in the embryo growth potential (EGP) and in the activity of two enzymes related to the weakening of the micropylar region of the endosperm (ME); endo-beta-mannanase and beta-mannosidase. The FR pulse also significantly stimulated the expression of DfGA3ox, a GA 3beta-hydroxylase, suggesting that the promotion of germination by the VLFR is associated with an increase in the synthesis of active gibberellins. The promotive action of the VLFR on germination is reduced when the FR pulse is immediately followed by a continuous FR treatment for 24 h (FRc). The effect of FRc cannot be reproduced by hourly FR pulses during the same period, showing that the antagonistic effect of FRc is a high-irradiance response (HIR). The action of the HIR in germination is associated with a decrease of both the mannan-degrading enzyme activity and the expression of DfMan in the ME, whereas no changes in the EGP were observed. The HIR also inhibits the accumulation of DfGA3ox in embryos, indicating that its action on germination is mediated, at least in part, through the modulation of active GA contents in seeds. This is the first report of a gene that participates in the VLFR-HIR antagonism in seeds.

Phytochrome control of the Arabidopsis transcriptome anticipates seedling exposure to light.

Phytochromes mediate a profound developmental shift when dark-grown seedlings are exposed to light. Here, we show that a subset of genes is upregulated in phytochrome B (phyB) mutants even before dark-grown Arabidopsis thaliana seedlings are exposed to light. Most of these genes bear the RY cis motif, which is a binding site of the transcription factor ABSCISIC ACID INSENSITIVE3 (ABI3), and the phyB mutation also enhances ABI3 expression. These changes in transcriptome have physiological consequences, because seedlings of the abi3 mutant showed enhanced responses to pulses of far-red light, whereas ABI3 overexpressers exhibited the opposite pattern. Seedlings of the wild type derived from seeds germinated in full darkness showed enhanced expression of genes bearing the RY cis motif and reduced responses to far-red light. We propose that, via changes in ABI3 expression, light, perceived mainly by phyB in the seed, generates a downstream transdevelopmental phase signal that preconditions the seedling to its most likely environment.

Increased phytochrome B alleviates density effects on tuber yield of field potato crops.

The possibility that reduced photomorphogenic responses could increase field crop yield has been suggested often, but experimental support is still lacking. Here, we report that ectopic expression of the Arabidopsis PHYB (phytochrome B) gene, a photoreceptor involved in detecting red to far-red light ratio associated with plant density, can increase tuber yield in field-grown transgenic potato (Solanum tuberosum) crops. Surprisingly, this effect was larger at very high densities, despite the intense reduction in the red to far-red light ratios and the concomitant narrowed differences in active phytochrome B levels between wild type and transgenics at these densities. Increased PHYB expression not only altered the ability of plants to respond to light signals, but they also modified the light environment itself. This combination resulted in larger effects of enhanced PHYB expression on tuber number and crop photosynthesis at high planting densities. The PHYB transgenics showed higher maximum photosynthesis in leaves of all strata of the canopy, and this effect was largely due to increased leaf stomatal conductance. We propose that enhanced PHYB expression could be used in breeding programs to shift optimum planting densities to higher levels.

The Cape Verde Islands allele of cryptochrome 2 enhances cotyledon unfolding in the absence of blue light in Arabidopsis.

We analyzed the natural genetic variation between Landsburg erecta (Ler) and Cape Verde Islands (Cvi) accessions by studying 105 recombinant inbred lines to search for players in the regulation of sensitivity to light signals perceived by phytochromes in etiolated seedlings of Arabidopsis. In seedlings grown under hourly pulses of far-red (FR) light, we identified three quantitative trait loci (QTLs; VLF3, VLF4, and VLF5) for hypocotyl growth inhibition and three different QTLs (VLF6, VLF7, and VLF1) for cotyledon unfolding. This indicates that different physiological outputs have selective regulation of sensitivity during de-etiolation. Ler alleles, compared with Cvi alleles, of VLF3, VLF4, VLF5, VLF7, and VLF1 enhanced, whereas the Ler allele of VLF6 reduced, the response to pulses of FR. We confirmed and narrowed down the position of some QTLs by using near-isogenic lines. VLF6 mapped close to the CRY2 (cryptochrome 2) gene. Transgenic Ler seedlings expressing the Cvi allele of CRY2 showed enhanced cotyledon unfolding under hourly pulses of FR compared with the wild type or transgenics expressing the CRY2-Ler allele. This response required phytochrome A. The cry1 cry2 double mutant lacking both cryptochromes showed reduced cotyledon unfolding under FR pulses. Because the CRY2-Cvi is a gain-of-function allele compared with CRY2-Ler, cryptochrome activity correlates positively with cotyledon unfolding under FR pulses. We conclude that the blue light photoreceptor cryptochrome 2 can modulate seedling photomorphogenesis in the absence of blue light. In addition to the nuclear loci, we identified cytoplasmic effects on seedling de-etiolation.

Expression analysis of a GA 20-oxidase in embryos from two sorghum lines with contrasting dormancy: possible participation of this gene in the hormonal control of germination.

The role of GAs in promoting seed germination is well known and experiments with seeds from different species have suggested the requirement of de novo synthesis of GAs upon imbibition for germination. There are also strong indications that the enhancement of GA synthesis is part of the mechanism through which environmental signals (i.e. light) induce germination. Since along the GA biosynthetic pathway, oxidation at C-20 carried out by GA 20-oxidases is thought to be a site of regulation, a cDNA clone encoding a GA 20-oxidase was isolated from embryos of sorghum (SbGA 20ox). Expression analysis of this gene in embryos within imbibed caryopses with low dormancy showed detectable amounts of the specific mRNA early upon incubation, increasing thereafter. In contrast, it remained barely detectable in embryos from dormant caryopses. Changes in endogenous GA4 levels were in agreement with those of SbGA 20ox mRNA, suggesting that GA production might be regulated differentially at the level of transcription of this gene. The expression of SbGA 20ox was enhanced in incubated embryos isolated from either type of caryopses, illustrating a physiological control exerted by the surrounding seed tissues on gene expression. The results also show that ABA leads to a suppression of transcription of this gene.

On the hormonal nature of the stimulatory effect of high incubation temperatures on germination of dormant sorghum (S. bicolor) caryopses.

• High incubation temperatures (i.e. 30°C) stimulate the germination of dormant sorghum grains. To test the hormonal nature of this response, experiments were carried out with two varieties with contrasting dormancy at harvest: Redland B2 (low dormancy, high germination percentages attained under a wide incubation thermal range) and IS 9530 (high dormancy, high germination percentages attained only at 30°C). • Redland B2 grains with reduced GA content (paclobutrazol-treated) reached high germination temperatures (c. 100%) only when incubated at 30°C. By contrast, IS 9530 grains with reduced ABA content (fluridone-treated) reached 100% germination at 30, 25, 20 and 15°C. • Incubation temperatures did not alter embryo responsiveness to ABA, nor did it modify the pattern of changes in embryo ABA content throughout incubation. Low GA3 concentrations (0.1 µm) were required to totally overcome the inhibition imposed by ABA in embryos incubated at 30°C; by contrast, even the highest GA3 concentrations used (1000 µm) were not able to revert ABA inhibition in embryos incubated at 15°C. • These results show the hormonal nature of the stimulatory effect of high incubation temperatures, and suggest that this effect is mediated by an increase in tissue responsiveness to GAs.

Effects of flood-influenced factors on seed germination of Ambrosia tenuifolia.

Strong fluctuations are exhibited by populations of the perennial herb Ambrosia tenuifolia in the grasslands of the Salado basin (Province of Buenos Aires, Argentina), an area frequently enduring prolonged floods. Flooding causes the death of most dicotyledon plants of the community, A. tenuifolia among them, opening numerous gaps of various sizes. After the recession of the flood the density of A. tenuifolia seedlings was higher in flooded than in non-flooded plots and it was larger in wider gaps. Canopy removal in non-flooded plots increased field seedling emergence of A. tenuifolia up to the levels found in flooded plots. Responses of the seeds in the soil to gap-associated environmental factors such as light quality and temperature regime were studied both in the field and under controlled were studied both in the field and under controlled conditions. Seedling emergence was significantly enhanced when the red:far-red ratio of natural light reaching the soil surface under the canopy of nonflooded plots was increased by means of copper sulfate filters. The influence of light quality and temperature on germination of the soil seed population was also tested using grassland soil monoliths or mesocosms, transported from the field to the laboratory, in which the canopy was clipped and the soil exposed to either red or far-red light and kept at constant or fluctuating temperatures. Significant seedling emergence was observed only when the soil samples were exposed to red light and incubated at alternating temperatures. No emergence was recorded in samples exposed to far-red light or incubated at a constant 25°C. Seeds stored dry in the laboratory were also stimulated to germinate by red light and alternating temperatures but only after dormancy was sufficiently decreased by low temperature stratification or by low temperature under immersion. The results are consistent with the hypothesis that primary dormancy of A. tenuifolia seeds is decreased by low temperatures in winter even if the seeds are submerged as happens when floods occur. The decrease in dormancy makes the seeds prone to be stimulated to germinate by the Pfr form of phytochrome in combination with alternating temperatures. These conditions are likely to be met in the gaps opened by the flood-caused death of dicotyledon plants.