Regulation of Root Exudation in Wheat Plants in Response to Alkali Stress.

Soil alkalization is an important environmental factor limiting crop production. Despite the importance of root secretion in the response of plants to alkali stress, the regulatory mechanism is unclear. In this study, we applied a widely targeted metabolomics approach using a local MS/MS data library constructed with authentic standards to identify and quantify root exudates of wheat under salt and alkali stresses. The regulatory mechanism of root secretion in alkali-stressed wheat plants was analyzed by determining transcriptional and metabolic responses. Our primary focus was alkali stress-induced secreted metabolites (AISMs) that showed a higher secretion rate in alkali-stressed plants than in control and salt-stressed plants. This secretion was mainly induced by high-pH stress. We discovered 55 AISMs containing -COOH groups, including 23 fatty acids, 4 amino acids, 1 amino acid derivative, 7 dipeptides, 5 organic acids, 9 phenolic acids, and 6 others. In the roots, we also discovered 29 metabolites with higher levels under alkali stress than under control and salt stress conditions, including 2 fatty acids, 3 amino acid derivatives, 1 dipeptide, 2 organic acids, and 11 phenolic acids. These alkali stress-induced accumulated carboxylic acids may support continuous root secretion during the response of wheat plants to alkali stress. In the roots, RNAseq analysis indicated that 5 6-phosphofructokinase (glycolysis rate-limiting enzyme) genes, 16 key fatty acid synthesis genes, and 122 phenolic acid synthesis genes have higher expression levels under alkali stress than under control and salt stress conditions. We propose that the secretion of multiple types of metabolites with a -COOH group is an important pH regulation strategy for alkali-stressed wheat plants. Enhanced glycolysis, fatty acid synthesis, and phenolic acid synthesis will provide more energy and substrates for root secretion during the response of wheat to alkali stress.

Chromosome translocation affects multiple phenotypes, causes genome-wide dysregulation of gene expression, and remodels metabolome in hexaploid wheat.

Chromosomal rearrangements (CRs) may occur in newly formed polyploids due to compromised meiotic fidelity. Moreover, CRs can be more readily tolerated in polyploids allowing their longer-term retention and hence potential spreading/fixation within a lineage. The direct functional consequences of CRs in plant polyploids remain unexplored. Here, we identified a heterozygous individual from a synthetic allohexaploid wheat in which the terminal parts of the long-arms of chromosomes 2D (approximately 193 Mb) and 4A (approximately 167 Mb) were reciprocally translocated. Five homogeneous translocation lines including both unbalanced and balanced types were developed by selfing fertilization of the founder mutant (RT [2DL; 4AL]-ter/1, reciprocal translocation). We investigated impacts of these translocations on phenotype, genome-wide gene expression and metabolome. We find that, compared with sibling wild-type, CRs in the form of both unbalanced and balanced translocations induced substantial changes of gene expression primarily via trans-regulation in the nascent allopolyploid wheat. The CRs also manifested clear phenotypic and metabolic consequences. In particular, the genetically balanced, stable reciprocal translocations lines showed immediate enhanced reproductive fitness relative to wild type. Our results underscore the profound impact of CRs on gene expression in nascent allopolyploids with wide-ranging phenotypic and metabolic consequences, suggesting CRs are an important source of genetic variation that can be exploited for crop breeding.

Homoeologous exchange enables rapid evolution of tolerance to salinity and hyper-osmotic stresses in a synthetic allotetraploid wheat.

The link between polyploidy and enhanced adaptation to environmental stresses could be a result of polyploidy itself harbouring higher tolerance to adverse conditions, or polyploidy possessing higher evolvability than diploids under stress conditions. Natural polyploids are inherently unsuitable to disentangle these two possibilities. Using selfed progenies of a synthetic allotetraploid wheat AT3 (AADD) along with its diploid parents, Triticum urartu TMU38 (AA) and Aegilops tauschii TQ27 (DD), we addressed the foregoing issue under abiotic salinity and hyper-osmotic (drought-like) stress. Under short duration of both stresses, euploid plants of AT3 showed intermediate tolerance of diploid parents; under life-long duration of both stresses, tolerant individuals to either stress emerged from selfed progenies of AT3, but not from comparable-sized diploid parent populations. Tolerance to both stresses were conditioned by the same two homoeologous exchanges (HEs; 2DS/2AS and 3DL/3AL), and at least one HE needed to be at the homozygous state. Transcriptomic analyses revealed that hyper-up-regulation of within-HE stress responsive genes of the A sub-genome origin is likely responsible for the dual-stress tolerant phenotypes. Our results suggest that HE-mediated inter-sub-genome rearrangements can be an important mechanism leading to adaptive evolution in allopolyploids as well as a promising target for genetic manipulation in crop improvement.

Extensive secretion of phenolic acids and fatty acids facilitates rhizosphere pH regulation in halophyte Puccinellia tenuiflora under alkali stress.

Puccinellia tenuiflora is a forage grass with high nutritional value that is an extreme alkali-tolerant halophyte: it can survive at pH 10-11. Root secretion is perceived as a major plant alkali tolerance mechanism. In the present study, we applied a widely targeted metabolomic approach to identify and quantify the root exudates of P. tenuiflora under alkali stress. We also surveyed the transcriptional and metabolic profiling of P. tenuiflora roots under salt (96-mM Na+ , pH 6.8) and alkali (96-mM Na+ , pH 9.6) stresses to reveal the biological processes mediating root secretion. In P. tenuiflora plants, 493 root exudates were detected under control conditions, 544 root exudates under salt stress conditions, and 607 root exudates under alkali stress conditions. Salt-stressed plants and alkali-stressed plants shared 64 root exudates, and 60 root exudates were unique to alkali-stressed plants. The secretion rate of 56 phenolic acids, 43 fatty acids, and 9 organic acids was faster in alkali-stressed roots than in control and salt-stressed roots. In P. tenuiflora roots, alkali stress enhanced the accumulation of 23 phenolic acids, five organic acids, and only one fatty acid. In addition, transcriptomic analysis revealed that alkali stress upregulated glycolysis and phenylpropanoid biosynthesis pathways in P. tenuiflora roots. Taken together, extensive secretion of phenolic acids and fatty acids promotes rhizosphere pH regulation of P. tenuiflora under alkali stress, which contributes to its strong alkali tolerance. The root secretion of P. tenuiflora upon alkali stress is highly organized. Enhanced glycolysis, phenylpropanoid biosynthesis, and organic acid synthesis in the roots provide more reducing power and carbon source for the root secretion process of alkali-stressed P. tenuiflora.

Salinity Tolerance in a Synthetic Allotetraploid Wheat (SlSlAA) Is Similar to Its Higher Tolerant Parent Aegilops longissima (SlSl) and Linked to Flavonoids Metabolism.

Allotetraploidization between A and S (closely related to B) genome species led to the speciation of allotetraploid wheat (genome BBAA). However, the immediate metabolic outcomes and adaptive changes caused by the allotetraploidization event are poorly understood. Here, we investigated how allotetraploidization affected salinity tolerance using a synthetic allotetraploid wheat line (genome SlSlAA, labeled as 4x), its Aegilops longissima (genome SlSl, labeled as SlSl) and Triticum urartu (AA genome, labeled as AA) parents. We found that the degree of salinity tolerance of 4x was similar to its SlSl parent, and both were substantially more tolerant to salinity stress than AA. This suggests that the SlSl subgenome exerts a dominant effect for this trait in 4x. Compared with SlSl and 4x, the salinity-stressed AA plants did not accumulate a higher concentration of Na+ in leaves, but showed severe membrane peroxidation and accumulated a higher concentration of ROS (H2O2 and O2 ⋅⁣-) and a lesser concentration of flavonoids, indicating that ROS metabolism plays a key role in saline sensitivity. Exogenous flavonoid application to roots of AA plants significantly relieved salinity-caused injury. Our results suggest that the higher accumulation of flavonoids in SlSl may contribute to ROS scavenging and salinity tolerance, and these physiological properties were stably inherited by the nascent allotetraploid SlSlAA.

Multiomics analysis provides insights into alkali stress tolerance of sunflower (Helianthus annuus L.).

Alkali stress is an extreme complex stress type, which exerts negative effects on plants via chemical destruction, osmotic stress, ion injury, nutrient deficiency, and oxygen deficiency. Soil alkalization has produced severe problems in some area, while plant alkali tolerance is poorly understood. Sunflower (Helianthus annuus L.) is an important oilseed crop with strong alkali tolerance. Here we exposed sunflower plants to alkali stress (NaHCO3/Na2CO3 = 9:1; pH 8.7) for whole life cycle. We applied transcriptomics, metabolomics, lipidomics and phytohormone analysis to elucidate the alkali tolerance mechanism of sunflower plant. Lipidomic analysis showed that alkali stress enhanced accumulation of saccharolipids and glycerolipids and lowered the accumulation of glycerophospholipids in sunflower seeds, indicating that alkali stress can change the lipid components of sunflower seeds, and that cultivating sunflower plants on alkalized farmlands will change the quality of sunflower seed oils. In addition, alkali stress downregulated expression of two rate-controlling genes of glycolysis in the leaves of sunflower but upregulated their expression in the roots. Enhanced glycolysis process provided more carbon sources and energy for alkali stress response of sunflower roots. Under alkali stress, accumulation of many fatty acids, amino acids, carbohydrates, and organic acids was greatly stimulated in sunflower roots. Alkali stress enhanced ACC, GA1, and ABA concentrations in the leaves but not in the roots, however, alkali stress elevated accumulation of BR (typhasterol) and CTK (Isopentenyladenosine) in the roots. We propose that multiple phytohormones and bioactive molecules interact to mediate alkali tolerance of sunflower.

Genome-wide DNA methylation analysis and biochemical responses provide insights into the initial domestication of halophyte Puccinellia tenuiflora.

KEY MESSAGE: Puccinellia tenuiflora was domesticated for two years by growing it under non-saline conditions, providing epigenetic and biochemical insights into the initial domestication of extreme halophytes. Some halophytes have economic value as crop species. The domestication of halophytes may offer hope in solving the problem of soil salinization. We domesticated a wild halophyte, Puccinellia tenuiflora, for two years by growing it under non-saline conditions in a greenhouse and used re-sequencing, genome-wide DNA methylation, biochemical, and transcriptome analyses to uncover the mechanisms underlying alterations in the halophyte's tolerance to saline following domestication. Our results showed that non-saline domestication altered the methylation status for a number of genes and transposable elements, resulting in a much higher frequency of hypomethylation than hypermethylation. These modifications to DNA methylation were observed in many critical salinity-tolerance genes, particularly their promoter regions or transcriptional start sites. Twenty-nine potassium channel genes were hypomethylated and three were hypermethylated, suggesting that the DNA methylation status of potassium channel genes was influenced by domestication. The accelerated uptake of potassium is a major salinity tolerance characteristic of P. tenuiflora. We propose that modifications to the DNA methylation of potassium channel genes may be associated with the development of salinity tolerance in this species. By assessing whether non-saline domestication could change the salinity tolerance of P. tenuiflora, we demonstrated that non-saline domesticated plants are less tolerant to saline, which may be attributable to altered sucrose metabolism. DNA methylation and transposable elements may, therefore, be integrated into an environment-sensitive molecular engine that promotes the rapid domestication of P. tenuiflora to enable its use as a crop plant.

Comparative Genomics and Transcriptomics of the Extreme Halophyte Puccinellia tenuiflora Provides Insights Into Salinity Tolerance Differentiation Between Halophytes and Glycophytes.

Halophytes and glycophytes exhibit clear differences in their tolerance to high levels of salinity. The genetic mechanisms underlying this differentiation, however, remain unclear. To unveil these mechanisms, we surveyed the evolution of salinity-tolerant gene families through comparative genomic analyses between the model halophyte Puccinellia tenuiflora and glycophytic Gramineae plants, and compared their transcriptional and physiological responses to salinity stress. Under salinity stress, the K+ concentration in the root was slightly enhanced in P. tenuiflora, but it was greatly reduced in the glycophytic Gramineae plants, which provided a physiological explanation for differences in salinity tolerance between P. tenuiflora and these glycophytes. Interestingly, several K+ uptake gene families from P. tenuiflora experienced family expansion and positive selection during evolutionary history. This gene family expansion and the elevated expression of K+ uptake genes accelerated K+ accumulation and decreased Na+ toxicity in P. tenuiflora roots under salinity stress. Positively selected P. tenuiflora K+ uptake genes may have evolved new functions that contributed to development of P. tenuiflora salinity tolerance. In addition, the expansion of the gene families involved in pentose phosphate pathway, sucrose biosynthesis, and flavonoid biosynthesis assisted the adaptation of P. tenuiflora to survival under high salinity conditions.

Adaptive strategy of allohexaploid wheat to long-term salinity stress.

BACKGROUND: Most studies of crop salinity tolerance are conducted under short-term stress condition within one growth stage. Understanding of the mechanisms of crop response to long-term salinity stress (LSS) is valuable for achieving the improvement of crop salinity tolerance. In the current study, we exposed allohexaploid wheat seeds to LSS conditions from germination stage to young seedling stage for 30 days. To elucidate the adaptive strategy of allohexaploid wheat to LSS, we analyzed chloroplast ultrastructure, leaf anatomy, transcriptomic profiling and concentrations of plant hormones and organic compatible solutes, comparing stressed and control plants. RESULTS: Transcriptomic profiling and biochemical analysis showed that energy partitioning between general metabolism maintenance and stress response may be crucial for survival of allohexaploid wheat under LSS. Under LSS, wheat appeared to shift energy from general maintenance to stress response through stimulating the abscisic acid (ABA) pathway and suppressing gibberellin and jasmonic acid pathways in the leaf. We further distinguished the expression status of the A, B, and D homeologs of any gene triad, and also surveyed the effects of LSS on homeolog expression bias for salinity-tolerant triads. We found that LSS had similar effects on expression of the three homeologs for most salinity-tolerant triads. However, in some of these triads, LSS induced different effects on the expression of the three homeologs. CONCLUSIONS: The shift of the energy from general maintenance to stress response may be important for wheat LSS tolerance. LSS influences homeolog expression bias of salinity-tolerant triads.

Genome of extreme halophyte Puccinellia tenuiflora.

BACKGROUND: Puccinellia tenuiflora, a forage grass, is considered a model halophyte given its strong tolerance for multiple stress conditions and its close genetic relationship with cereals. This halophyte has enormous values for improving our understanding of salinity tolerance mechanisms. The genetic information of P. tenuiflora also is a potential resource that can be used for improving the salinity tolerance of cereals. RESULTS: Here, we sequenced and assembled the P. tenuiflora genome (2n = 14) through the combined strategy of Illumina, PacBio, and 10× genomic technique. We generated 43.2× PacBio long reads, 123.87× 10× genomic reads, and 312.6× Illumina reads. Finally, we assembled 2638 scaffolds with a total size of 1.107 Gb, contig N50 of 117 kb, and scaffold N50 of 950 kb. We predicted 39,725 protein-coding genes, and identified 692 tRNAs, 68 rRNAs, 702 snRNAs, 1376 microRNAs, and 691 Mb transposable elements. CONCLUSIONS: We deposited the genome sequence in NCBI and the Genome Warehouse in National Genomics Data Center. Our work may improve current understanding of plant salinity tolerance, and provides extensive genetic resources necessary for improving the salinity and drought tolerance of cereals.

An extracted tetraploid wheat harbouring the BBAA component of common wheat shows anomalous shikimate and sucrose metabolism.

BACKGROUND: The BBAA subgenomes of hexaploid common wheat are structurally intact, which makes it possible to extract the BBAA subgenomes to constitute a novel plant type, namely, extracted tetraploid wheat (ETW). ETW displays multiple abnormal phenotypes such as massively reduced biomass and abnormal spike development, compared to extant tetraploid wheat with a BBAA genome. The genetic, biochemical and physiological basis underlying the phenotypic abnormality of ETW remains unknown. RESULTS: To explore the biochemical basis of these phenotypic abnormalities, we analysed the metabolomic and proteomic profiles and quantified 46 physiological traits of ETW in comparison with its common wheat donor (genome BBAADD), and a durum tetraploid wheat cultivar (genome BBAA). Among these three types of wheat, ETW showed a saliently different pattern of nutrient accumulation and seed quality, markedly lower concentrations of many metabolites involved in carbohydrate metabolism, and higher concentrations of many metabolites related to amino acids. Among the metabolites, changes in shikimate and sucrose were the most conspicuous. Higher levels of shikimate and lower levels of sucrose influence many metabolic processes including carbohydrate and amino acid metabolism, which may contribute to the phenotypic abnormalities. Gene expression assay showed downregulation of a shikimate degradation enzyme (5-enolpyruvylshikimate-3-phosphate synthase) coding gene and upregulation of several genes coding for the sucrose hydrolysis enzyme, which could explain the higher levels of shikimate and lower levels of sucrose, respectively. CONCLUSIONS: Our results suggest that significant and irreversible biochemical changes have occurred in the BBAA subgenomes of common wheat during the course of its co-evolution with the DD subgenome at the hexaploid level.

Proteomic profiling sheds light on alkali tolerance of common wheat (Triticum aestivum L.).

Alkali (high-pH) stress is an important factor limiting agricultural production and has complex effects on plant metabolism. Transcriptomics is widely used in the discovery of stress-response genes, but it provides only a rough estimation for gene expression. Proteomics may be more helpful than transcriptomics for the discovery and identification of stress-response genes. In this study, wheat plants were treated with sodic alkaline stress (50 mM, NaHCO3: Na2CO3 = 1:1; pH 9.7), and then proteomic analysis was carried out on control and stressed plants. We detected 3,104 proteins, including 69 alkaline stress-response proteins. Five superoxide dismutases, three malate dehydrogenases, three dehydrin proteins, and one V-ATPase protein were upregulated in sodic alkaline-stressed wheat roots. We propose that these salinity response proteins may be important for ion homeostasis and osmotic regulation of sodic alkaline-stressed wheat. Additionally, two malic enzymes and many enzymes involved in the tricarboxylic acid cycle (TCA) were downregulated in the roots. The upregulation of malate dehydrogenase and the downregulation of TCA enzymes and malic enzymes may enhance the accumulation of malate in sodic alkaline-stressed wheat roots. Previous studies have demonstrated that the accumulation of malate in roots is a crucial adaptive mechanism of wheat to sodic alkaline stress. Herein, our proteomics results provided molecular insights into this adaptive mechanism.

Rapid and pervasive development- and tissue-specific homeolog expression partitioning in newly formed inter-subspecific rice segmental allotetraploids.

BACKGROUND: In diverse plant taxa, whole-genome duplication (WGD) events are major sources of phenotypic novelty. Studies of gene expression in synthetic polyploids have shown immediate expression and functional partitioning of duplicated genes among different tissues. Many studies of the tissue-specific homeolog expression partitioning have focused on allopolyploids that have very different parental genomes, while few studies have focused on autopolyploids or allopolyploids that have similar parental genomes. RESULTS: In this study, we used a set of reciprocal F1 hybrids and synthetic tetraploids constructed from subspecies (japonica and indica) of Asian rice (Oryza sativa L.) as a model to gain insights into the expression partitioning of homeologs among tissues in a developmental context. We assayed the tissue-specific silencing (TSS) of the parental homeologs of 30 key genes in the hybrids and tetraploids relative to the in vitro "hybrids" (parental mixes) using Sequenom MassARRAY. We found that the parental mix and synthetic tetraploids had higher frequencies of homeolog TSS than the F1, revealing an instantaneous role of WGD on homeolog expression partitioning. CONCLUSIONS: Our observations contradicted those of previous studies in which newly formed allopolyploids had a low TSS frequency, similar to that of F1 hybrids, suggesting that the impact of WGD on homeolog expression requires a longer time to manifest. In addition, we found that the TSS frequency in the tetraploids varied at different growth stages and that roots had a much higher frequency of TSS than leaves, which indicated that developmental and metabolic traits may influence the expression states of duplicated genes in newly formed plant polyploids.

A newly formed hexaploid wheat exhibits immediate higher tolerance to nitrogen-deficiency than its parental lines.

BACKGROUND: It is known that hexaploid common wheat (Triticum aestivum L.) has stronger adaptability to many stressful environments than its tetraploid wheat progenitor. However, the physiological basis and evolutionary course to acquire these enhanced adaptabilities by common wheat remain understudied. Here, we aimed to investigate whether and by what means tolerance to low-nitrogen manifested by common wheat may emerge immediately following allohexaploidization. RESULTS: We compared traits related to nitrogen (N) metabolism in a synthetic allohexaploid wheat (neo-6×, BBAADD) mimicking natural common wheat, together with its tetraploid (BBAA, 4×) and diploid (DD, 2×) parents. We found that, under low nitrogen condition, neo-6× maintained largely normal photosynthesis, higher shoot N accumulation, and better N assimilation than its 4× and 2× parents. We showed that multiple mechanisms underlie the enhanced tolerance to N-deficiency in neo-6×. At morphological level, neo-6× has higher root/shoot ratio of biomass than its parents, which might be an adaptive growth strategy as more roots feed less shoots with N, thereby enabling higher N accumulation in the shoots. At electrophysiological level, H+ efflux in neo-6× is higher than in its 4× parent. A stronger H+ efflux may enable a higher N uptake capacity of neo-6×. At gene expression level, neo-6× displayed markedly higher expression levels of critical genes involved in N uptake than both of its 4× and 2× parents. CONCLUSIONS: This study documents that allohexaploid wheat can attain immediate higher tolerance to N-deficiency compared with both of its 4× and 2× parents, and which was accomplished via multiple mechanisms.

Meiotic chromosome stability of a newly formed allohexaploid wheat is facilitated by selection under abiotic stress as a spandrel.

Polyploidy is a prominent route to speciation in plants; however, this entails resolving the challenges of meiotic instability facing abrupt doubling of chromosome complement. This issue remains poorly understood. We subjected progenies of a synthetic hexaploid wheat, analogous to natural common wheat, but exhibiting extensive meiotic chromosome instability, to heat or salt stress. We selected stress-tolerant cohorts and generated their progenies under normal condition. We conducted fluorescent in situ hybridization/genomic in situ hybridization-based meiotic/mitotic analysis, RNA-Seq and virus-induced gene silencing (VIGS)-mediated assay of meiosis candidate genes. We show that heritability of stress tolerance concurred with increased euploidy frequency due to enhanced meiosis stability. We identified a set of candidate meiosis genes with altered expression in the stress-tolerant plants vs control, but the expression was similar to that of common wheat (cv Chinese Spring, CS). We demonstrate VIGS-mediated downregulation of individual candidate meiosis genes in CS is sufficient to confer an unstable meiosis phenotype mimicking the synthetic wheat. Our results suggest that heritable regulatory changes of preexisting meiosis genes may be hitchhiked as a spandrel of stress tolerance, which significantly improves meiosis stability in the synthetic wheat. Our findings implicate a plausible scenario that the meiosis machinery in hexaploid wheat may have already started to evolve at its onset stage.

Transcriptomic Profiling and Physiological Responses of Halophyte Kochia sieversiana Provide Insights into Salt Tolerance.

Halophytes are remarkable plants that can tolerate extremely high-salinity conditions, and have different salinity tolerance mechanisms from those of glycophytic plants. In this work, we investigated the mechanisms of salinity tolerance of an extreme halophyte, Kochia sieversiana (Pall.) C. A. M, using RNA sequencing and physiological tests. The results showed that moderate salinity stimulated the growth and water uptake of K. sieversiana and, even under 480-mM salinity condition, K. sieversiana maintained an extremely high water content. This high water content may be a specific adaptive strategy of K. sieversiana to high salinity. The physiological analysis indicated that increasing succulence and great accumulations of sodium, alanine, sucrose, and maltose may be favorable to the water uptake and osmotic regulation of K. sieversiana under high-salinity stress. Transcriptome data indicated that some aquaporin genes and potassium (K+) transporter genes may be important for water uptake and ion balance, respectively, while different members of those gene families were employed under low- and high-salinity stresses. In addition, several aquaporin genes were up-regulated in low- but not high-salinity stressed roots. The highly expressed aquaporin genes may allow low-salinity stressed K. sieversiana plants to uptake more water than control plants. The leaf K+/root K+ ratio was enhanced under low- but not high-salinity stress, which suggested that low salinity might promote K+ transport from the roots to the shoots. Hence, we speculated that low salinity might allow K. sieversiana to uptake more water and transport more K+ from roots to shoots, increasing the growth rate of K. sieversiana.

Segmental allotetraploidy generates extensive homoeologous expression rewiring and phenotypic diversity at the population level in rice.

Allopolyploidization, that is, concomitant merging and doubling of two or more divergent genomes in a common nucleus/cytoplasm, is known to instantly alter genomewide transcriptome dynamics, a phenomenon referred to as "transcriptomic shock." However, the immediate effects of transcriptomic alteration in generating phenotypic diversity at the population level remain underinvestigated. Here, we employed the MassARRAY-based Sequenom platform to assess and compare orthologous, allelic and homoeologous gene expression status in two tissues (leaf and root) of a set of randomly chosen individuals from populations of parental rice subspecies (indica and japonica), in vitro "hybrids" (parental mixes), reciprocal F1 hybrids and reciprocal tetraploids at the 5th-selfed generation (S5). We show that hybridization and whole genome duplication (WGD) have opposing effects on allelic and homoeologous expression in the F1 hybrids and tetraploids, respectively. Whereas hybridization exerts strong attenuating effects on allelic expression differences in diploid hybrids, WGD augments the intrinsic parental differences and generates extensive and variable homoeolog content which triggers diversification in expression patterning among the tetraploid plants. Coupled with the vast phenotypic diversity observed among the tetraploid individuals, our results provide experimental evidence in support of the notion that allopolyploidy catalyses rapid phenotypic diversification in higher plants. Our data further suggest that largely stochastic homoeolog content reshuffling rather than alteration in total expression level may be an important feature of evolution in young segmental allopolyploids, which underlies rapid expression diversity at the population level.

Comparison of Ionomic and Metabolites Response under Alkali Stress in Old and Young Leaves of Cotton (Gossypium hirsutum L.) Seedlings.

Soil salinization is an important agriculture-related environmental problem. Alkali stress and salt stress strongly influence the metabolic balance in plants. Salt and alkali stresses exert varied effects on old and young tissues, which display different adaptive strategies. In this study, we used cotton (Gossypium hirsutum L.) plants as experimental material to investigate whether alkali stress induces ionic and metabolism changes in old and young leaves of cotton plants exposed to alkali stress. Results showed that alkali stress exerted a considerably stronger growth inhibition on old leaves than on young leaves. Under alkali stress, young leaves can maintain low Na and high K contents and retain relatively stable tricarboxylic acid cycle, resulting in greater accumulation of photosynthetic metabolites. In terms of metabolic response, the young and old leaves clearly displayed different mechanisms of osmotic regulation. The amounts of inositol and mannose significantly increased in both old and young leaves of cotton exposed to alkali stress, and the extent of increase was higher in young leaves than in old leaves. In old leaves, synthesis of amino acids, such as GABA, valine, and serine, was dramatically enhanced, and this phenomenon is favorable for osmotic adjustment and membrane stability. Organs at different developmental stages possibly display different mechanisms of metabolic regulation under stress condition. Thus, we propose that future investigations on alkali stress should use more organs obtained at different developmental stages.

Evolution of physiological responses to salt stress in hexaploid wheat.

Hexaploid bread wheat (Triticum aestivum L., genome BBAADD) is generally more salt tolerant than its tetraploid wheat progenitor (Triticum turgidum L.). However, little is known about the physiological basis of this trait or about the relative contributions of allohexaploidization and subsequent evolutionary genetic changes on the trait development. Here, we compared the salt tolerance of a synthetic allohexaploid wheat (neo-6x) with its tetraploid (T. turgidum; BBAA) and diploid (Aegilops tauschii; DD) parents, as well as a natural hexaploid bread wheat (nat-6x). We studied 92 morphophysiological traits and analyzed homeologous gene expression of a major salt-tolerance gene High-Affinity K(+) Transporter 1;5 (HKT1;5). We observed that under salt stress, neo-6x exhibited higher fitness than both of its parental genotypes due to inheritance of favorable traits like higher germination rate from the 4x parent and the stronger root Na(+) retention capacity from the 2x parent. Moreover, expression of the D-subgenome HKT1;5 homeolog, which is responsible for Na(+) removal from the xylem vessels, showed an immediate transcriptional reprogramming following allohexaploidization, i.e., from constitutive high basal expression in Ae. tauschii (2x) to salt-induced expression in neo-6x. This phenomenon was also witnessed in the nat-6x. An integrated analysis of 92 traits showed that, under salt-stress conditions, neo-6x resembled more closely the 2x than the 4x parent, suggesting that the salt stress induces enhanced expressivity of the D-subgenome homeologs in the synthetic hexaploid wheat. Collectively, the results suggest that condition-dependent functionalization of the subgenomes might have contributed to the wide-ranging adaptability of natural hexaploid wheat.

Evolution of the BBAA component of bread wheat during its history at the allohexaploid level.

Subgenome integrity in bread wheat (Triticum aestivum; BBAADD) makes possible the extraction of its BBAA component to restitute a novel plant type. The availability of such a ploidy-reversed wheat (extracted tetraploid wheat [ETW]) provides a unique opportunity to address whether and to what extent the BBAA component of bread wheat has been modified in phenotype, karyotype, and gene expression during its evolutionary history at the allohexaploid level. We report here that ETW was anomalous in multiple phenotypic traits but maintained a stable karyotype. Microarray-based transcriptome profiling identified a large number of differentially expressed genes between ETW and natural tetraploid wheat (Triticum turgidum), and the ETW-downregulated genes were enriched for distinct Gene Ontology categories. Quantitative RT-PCR analysis showed that gene expression differences between ETW and a set of diverse durum wheat (T. turgidum subsp durum) cultivars were distinct from those characterizing tetraploid cultivars per se. Pyrosequencing revealed that the expression alterations may occur to either only one or both of the B and A homoeolog transcripts in ETW. A majority of the genes showed additive expression in a resynthesized allohexaploid wheat. Analysis of a synthetic allohexaploid wheat and diverse bread wheat cultivars revealed the rapid occurrence of expression changes to the BBAA subgenomes subsequent to allohexaploidization and their evolutionary persistence.