The dimorphic diaspore model Aethionema arabicum (Brassicaceae): Distinct molecular and morphological control of responses to parental and germination temperatures.

Plants in habitats with unpredictable conditions often have diversified bet-hedging strategies that ensure fitness over a wider range of variable environmental factors. A striking example is the diaspore (seed and fruit) heteromorphism that evolved to maximize species survival in Aethionema arabicum (Brassicaceae) in which external and endogenous triggers allow the production of two distinct diaspores on the same plant. Using this dimorphic diaspore model, we identified contrasting molecular, biophysical, and ecophysiological mechanisms in the germination responses to different temperatures of the mucilaginous seeds (M+ seed morphs), the dispersed indehiscent fruits (IND fruit morphs), and the bare non-mucilaginous M- seeds obtained by pericarp (fruit coat) removal from IND fruits. Large-scale comparative transcriptome and hormone analyses of M+ seeds, IND fruits, and M- seeds provided comprehensive datasets for their distinct thermal responses. Morph-specific differences in co-expressed gene modules in seeds, as well as in seed and pericarp hormone contents, identified a role of the IND pericarp in imposing coat dormancy by generating hypoxia affecting abscisic acid (ABA) sensitivity. This involved expression of morph-specific transcription factors, hypoxia response, and cell wall remodeling genes, as well as altered ABA metabolism, transport, and signaling. Parental temperature affected ABA contents and ABA-related gene expression and altered IND pericarp biomechanical properties. Elucidating the molecular framework underlying the diaspore heteromorphism can provide insight into developmental responses to globally changing temperatures.

A tale of two morphs: developmental patterns and mechanisms of seed coat differentiation in the dimorphic diaspore model Aethionema arabicum (Brassicaceae).

The developmental transition from a fertilized ovule to a dispersed diaspore (seed or fruit) involves complex differentiation processes of the ovule's integuments leading to the diversity in mature seed coat structures in angiosperms. In this study, comparative imaging and transcriptome analysis were combined to investigate the morph-specific developmental differences during outer seed coat differentiation and mucilage production in Aethionema arabicum, the Brassicaceae model for diaspore dimorphism. One of the intriguing adaptations of this species is the production and dispersal of morphologically distinct, mucilaginous and non-mucilaginous diaspores from the same plant (dimorphism). The dehiscent fruit morph programme producing multiple mucilaginous seed diaspores was used as the default trait combination, similar to Arabidopsis thaliana, and was compared with the indehiscent fruit morph programme leading to non-mucilaginous diaspores. Synchrotron-based radiation X-ray tomographic microscopy revealed a co-ordinated framework of morph-specific early changes in internal anatomy of developing A. arabicum gynoecia including seed abortion in the indehiscent programme and mucilage production by the mucilaginous seed coat. The associated comparative analysis of the gene expression patterns revealed that the unique seed coat dimorphism of Ae. arabicum provides an excellent model system for comparative study of the control of epidermal cell differentiation and mucilage biosynthesis by the mucilage transcription factor cascade and their downstream cell wall and mucilage remodelling genes. Elucidating the underlying molecular framework of the dimorphic diaspore syndrome is key to understanding differential regulation of bet-hedging survival strategies in challenging environments, timely in the face of global climatic change.

Usability of reference-free transcriptome assemblies for detection of differential expression: a case study on Aethionema arabicum dimorphic seeds.

BACKGROUND: RNA-sequencing analysis is increasingly utilized to study gene expression in non-model organisms without sequenced genomes. Aethionema arabicum (Brassicaceae) exhibits seed dimorphism as a bet-hedging strategy - producing both a less dormant mucilaginous (M+) seed morph and a more dormant non-mucilaginous (NM) seed morph. Here, we compared de novo and reference-genome based transcriptome assemblies to investigate Ae. arabicum seed dimorphism and to evaluate the reference-free versus -dependent approach for identifying differentially expressed genes (DEGs). RESULTS: A de novo transcriptome assembly was generated using sequences from M+ and NM Ae. arabicum dry seed morphs. The transcripts of the de novo assembly contained 63.1% complete Benchmarking Universal Single-Copy Orthologs (BUSCO) compared to 90.9% for the transcripts of the reference genome. DEG detection used the strict consensus of three methods (DESeq2, edgeR and NOISeq). Only 37% of 1533 differentially expressed de novo assembled transcripts paired with 1876 genome-derived DEGs. Gene Ontology (GO) terms distinguished the seed morphs: the terms translation and nucleosome assembly were overrepresented in DEGs higher in abundance in M+ dry seeds, whereas terms related to mRNA processing and transcription were overrepresented in DEGs higher in abundance in NM dry seeds. DEGs amongst these GO terms included ribosomal proteins and histones (higher in M+), RNA polymerase II subunits and related transcription and elongation factors (higher in NM). Expression of the inferred DEGs and other genes associated with seed maturation (e.g. those encoding late embryogenesis abundant proteins and transcription factors regulating seed development and maturation such as ABI3, FUS3, LEC1 and WRI1 homologs) were put in context with Arabidopsis thaliana seed maturation and indicated that M+ seeds may desiccate and mature faster than NM. The 1901 transcriptomic DEG set GO-terms had almost 90% overlap with the 2191 genome-derived DEG GO-terms. CONCLUSIONS: Whilst there was only modest overlap of DEGs identified in reference-free versus -dependent approaches, the resulting GO analysis was concordant in both approaches. The identified differences in dry seed transcriptomes suggest mechanisms underpinning previously identified contrasts between morphology and germination behaviour of M+ and NM seeds.

Dispersal biophysics and adaptive significance of dimorphic diaspores in the annual Aethionema arabicum (Brassicaceae).

Heteromorphic diaspores (fruits and seeds) are an adaptive bet-hedging strategy to cope with spatiotemporally variable environments, particularly fluctuations in favourable temperatures and unpredictable precipitation regimes in arid climates. We conducted comparative analyses of the biophysical and ecophysiological properties of the two distinct diaspores (mucilaginous seed (M+ ) vs indehiscent (IND) fruit) in the dimorphic annual Aethionema arabicum (Brassicaceae), linking fruit biomechanics, dispersal aerodynamics, pericarp-imposed dormancy, diaspore abscisic acid (ABA) concentration, and phenotypic plasticity of dimorphic diaspore production to its natural habitat and climate. Two very contrasting dispersal mechanisms of the A. arabicum dimorphic diaspores were revealed. Dehiscence of large fruits leads to the release of M+ seed diaspores, which adhere to substrata via seed coat mucilage, thereby preventing dispersal (antitelechory). IND fruit diaspores (containing nonmucilaginous seeds) disperse by wind or water currents, promoting dispersal (telechory) over a longer range. The pericarp properties confer enhanced dispersal ability and degree of dormancy on the IND fruit morph to support telechory, while the M+ seed morph supports antitelechory. Combined with the phenotypic plasticity to produce more IND fruit diaspores in colder temperatures, this constitutes a bet-hedging survival strategy to magnify the prevalence in response to selection pressures acting over hilly terrain.

Aethionema arabicum: a novel model plant to study the light control of seed germination.

The timing of seed germination is crucial for seed plants and is coordinated by internal and external cues, reflecting adaptations to different habitats. Physiological and molecular studies with lettuce and Arabidopsis thaliana have documented a strict requirement for light to initiate germination and identified many receptors, signaling cascades, and hormonal control elements. In contrast, seed germination in several other plants is inhibited by light, but the molecular basis of this alternative response is unknown. We describe Aethionema arabicum (Brassicaceae) as a suitable model plant to investigate the mechanism of germination inhibition by light, as this species has accessions with natural variation between light-sensitive and light-neutral responses. Inhibition of germination occurs in red, blue, or far-red light and increases with light intensity and duration. Gibberellins and abscisic acid are involved in the control of germination, as in Arabidopsis, but transcriptome comparisons of light- and dark-exposed A. arabicum seeds revealed that, upon light exposure, the expression of genes for key regulators undergo converse changes, resulting in antipodal hormone regulation. These findings illustrate that similar modular components of a pathway in light-inhibited, light-neutral, and light-requiring germination among the Brassicaceae have been assembled in the course of evolution to produce divergent pathways, likely as adaptive traits.

Aethionema arabicum dimorphic seed trait resetting during transition to seedlings.

The transition from germinating seeds to emerging seedlings is one of the most vulnerable plant life cycle stages. Heteromorphic diaspores (seed and fruit dispersal units) are an adaptive bet-hedging strategy to cope with spatiotemporally variable environments. While the roles and mechanisms of seedling traits have been studied in monomorphic species, which produce one type of diaspore, very little is known about seedlings in heteromorphic species. Using the dimorphic diaspore model Aethionema arabicum (Brassicaceae), we identified contrasting mechanisms in the germination responses to different temperatures of the mucilaginous seeds (M+ seed morphs), the dispersed indehiscent fruits (IND fruit morphs), and the bare non-mucilaginous M- seeds obtained from IND fruits by pericarp (fruit coat) removal. What follows the completion of germination is the pre-emergence seedling growth phase, which we investigated by comparative growth assays of early seedlings derived from the M+ seeds, bare M- seeds, and IND fruits. The dimorphic seedlings derived from M+ and M- seeds did not differ in their responses to ambient temperature and water potential. The phenotype of seedlings derived from IND fruits differed in that they had bent hypocotyls and their shoot and root growth was slower, but the biomechanical hypocotyl properties of 15-day-old seedlings did not differ between seedlings derived from germinated M+ seeds, M- seeds, or IND fruits. Comparison of the transcriptomes of the natural dimorphic diaspores, M+ seeds and IND fruits, identified 2,682 differentially expressed genes (DEGs) during late germination. During the subsequent 3 days of seedling pre-emergence growth, the number of DEGs was reduced 10-fold to 277 root DEGs and 16-fold to 164 shoot DEGs. Among the DEGs in early seedlings were hormonal regulators, in particular for auxin, ethylene, and gibberellins. Furthermore, DEGs were identified for water and ion transporters, nitrate transporter and assimilation enzymes, and cell wall remodeling protein genes encoding enzymes targeting xyloglucan and pectin. We conclude that the transcriptomes of seedlings derived from the dimorphic diaspores, M+ seeds and IND fruits, undergo transcriptional resetting during the post-germination pre-emergence growth transition phase from germinated diaspores to growing seedlings.

Survey of artemisinin production by diverse Artemisia species in northern Pakistan.

BACKGROUND: Artemisinin is the current drug of choice for treatment of malaria and a number of other diseases. It is obtained from the annual herb, Artemisia annua and some microbial sources by genetic engineering. There is a great concern that the artemisinin production at current rate will not meet the increasing demand by the pharmaceutical industry, so looking for additional sources is imperative. METHODS: In current study, artemisinin concentration was analysed and compared in the flowers, leaves, roots and stems of Artemisia annua and 14 other Artemisia species including two varieties each for Artemisia roxburghiana and Artemisia dracunculus using high performance liquid chromatography (HPLC). RESULTS: The highest artemisinin concentration was detected in the leaves (0.44 ± 0.03%) and flowers (0.42 ± 0.03%) of A. annua, followed by the flowers (0.34 ± .02%) of A. bushriences and leaves (0.27 ± 0%) of A. dracunculus var dracunculus. The average concentration of artemisinin varied in the order of flowers > leaves > stems > roots. CONCLUSION: This study identifies twelve novel plant sources of artemisinin, which may be helpful for pharmaceutical production of artemisinin. This is the first report of quantitative comparison of artemisinin among a large number of Artemisia species.

Nutrient homeostasis within the plant circadian network.

Circadian clocks have evolved to enhance adaptive physiology in the predictable, fluctuating environment caused by the rotation of the planet. Nutrient acquisition is central to plant growth performance and the nutrient demands of a plant change according to the time of day. Therefore, major aspects of nutrient homeostasis, including carbon assimilation and mineral uptake, are under circadian control. It is also emerging that there is feedback of nutritional status to the circadian clock to integrate these processes. This review will highlight recent insights into the role of the circadian clock in regulating plant nutrition as well as discuss the role for nutrients in affecting circadian function.

Agrobacterium-mediated transformation of tomato with rolB gene results in enhancement of fruit quality and foliar resistance against fungal pathogens.

Tomato (Solanum lycopersicum L.) is the second most important cultivated crop next to potato, worldwide. Tomato serves as an important source of antioxidants in human diet. Alternaria solani and Fusarium oxysporum cause early blight and vascular wilt of tomato, respectively, resulting in severe crop losses. The foremost objective of the present study was to generate transgenic tomato plants with rolB gene and evaluate its effect on plant morphology, nutritional contents, yield and resistance against fungal infection. Tomato cv. Rio Grande was transformed via Agrobacterium tumefaciens harbouring rolB gene of Agrobacterium rhizogenes. rolB. Biochemical analyses showed considerable improvement in nutritional quality of transgenic tomato fruits as indicated by 62% increase in lycopene content, 225% in ascorbic acid content, 58% in total phenolics and 26% in free radical scavenging activity. Furthermore, rolB gene significantly improved the defence response of leaves of transgenic plants against two pathogenic fungal strains A. solani and F. oxysporum. Contrarily, transformed plants exhibited altered morphology and reduced fruit yield. In conclusion, rolB gene from A. rhizogenes can be used to generate transgenic tomato with increased nutritional contents of fruits as well as improved foliar tolerance against fungal pathogens.

Effects of vegetative and flowering stages on the biosynthesis of artemisinin in Artemisia species.

Artemisinin is an endoperoxide sesquiterpene lactone, and has been proven to be very effective in treating drug resistant cases of malaria, cancer, etc. The compound is obtained from Artemisia species. In the current study, the effects of vegetative and flowering stages on artemisinin production were studied, to determine the proper harvesting time of naturally growing Artemisia species with the highest levels of artemisinin. Eight Artemisia species along with two varieties were selected for this analytical work. The results showed that artemisinin content was high in the leaves of Artemisia indica, A. sieversiana, A. roxburghiana var. roxburghiana, A. roxburghiana var. gratae, and A. parviflora at the flowering stage. The highest artemisinin content was measured in the leaves of A. dracunculus var. dracunculus. Upon comparisons of artemisinin content among the individual plant species, the highest amount of artemisinin was again in A. dracunculus var. dracunculus followed by A. sieversiana when harvested at the flowering stage. In overall comparisons, the plants at the flowering stage showed high levels of artemisinin, which is deemed the optimum harvesting time of Artemisia species in Pakistan for maximum artemisinin content.