Comparative metabolomic profiling of Arabidopsis thaliana roots and leaves reveals complex response mechanisms induced by a seaweed extract.

Seaweed extracts are a prominent class of biostimulants that enhance plant health and tolerance to biotic and abiotic stresses due to their unique bioactive components. However, the mechanisms of action of biostimulants are still unknown. Here, we have used a metabolomic approach, a UHPLC-MS method, to uncover the mechanisms induced following application to Arabidopsis thaliana of a seaweed extract derived from Durvillaea potatorum and Ascophyllum nodosum. We have identified, following the application of the extract, key metabolites and systemic responses in roots and leaves across 3 timepoints (0, 3, 5 days). Significant alterations in metabolite accumulation or reduction were found for those belonging to broad groups of compounds such as lipids, amino acids, and phytohormones; and secondary metabolites such as phenylpropanoids, glucosinolates, and organic acids. Strong accumulations of TCA cycle and N-containing and defensive metabolites such as glucosinolates were also found revealing the enhancement of carbon and nitrogen metabolism and defence systems. Our study has demonstrated that application of seaweed extract dramatically altered the metabolomic profiles of Arabidopsis and revealed differences in roots and leaves that varied across the timepoints tested. We also show clear evidence of systemic responses that were initiated in the roots and resulted in metabolic alterations in the leaves. Collectively, our results suggest that this seaweed extract promotes plant growth and activates defence systems by altering various physiological processes at the individual metabolite level.

Effect of a Fortified Biostimulant Extract on Tomato Plant Productivity, Physiology, and Growing Media Properties.

The pursuit of sustainable and productive agriculture demands the exploration of innovative approaches to improve plant productivity and soil health. The utilization of natural agricultural biostimulants, such as extracts from seaweed, fish, and humus, has gained prominence as an ecological strategy to achieve this goal. In this study we investigated the effectiveness of a fortified biostimulant extract (FBE), composed of extracts from seaweed, fish, and humus, on tomato plant physiology, productivity, and growing media properties, and estimated carbon emissions associated with tomato production. The FBE was applied to the growing media of tomato plants produced in a greenhouse, in experiments over two growing seasons. The productivity assessments demonstrated that the application of FBE significantly increased tomato fruit yield by 20% and relative marketable fruit yield by 27%, and reduced estimated greenhouse gas (GHG) emissions associated with production by 29%. FBE treatment improved plant shoot and root biomass, accelerated flower and fruit set initiation, and increased chlorophyll content in leaves, resulting in enhanced plant physiology and advanced development. FBE treatment positively influenced the availability of crucial nutrients such as nitrogen, phosphorus, and iron in the growing media. FBE promoted the growth of total active microbes in the growing media, particularly the fungal population, which plays an important role in nutrient cycling and health. These findings highlight the beneficial effects of the FBE due to enhanced plant productivity and growth, improved fertility, the promotion of beneficial plant and growing media interactions, and the reduction in estimated GHG emissions.

Seaweed Extract-Stimulated Priming in Arabidopsis thaliana and Solanum lycopersicum.

Plant priming is an induced physiological state where plants are protected from biotic and abiotic stresses. Whether seaweed extracts promote priming is largely unknown as is the mechanism by which priming may occur. In this study, we examined the effect of a seaweed extract (SWE) on two distinct stages of plant priming (priming phase and post-challenge primed state) by characterising (i) plant gene expression responses using qRT-PCR and (ii) signal transduction responses by evaluating reactive oxygen species (ROS) production. The SWE is made from the brown algae Ascophyllum nodosum and Durvillaea potatorum. The priming phase was examined using both Arabidopsis thaliana and Solanum lycopersicum. At this stage, the SWE up-regulated key priming-related genes, such as those related to systemic acquired resistance (SAR) and activated the production of ROS. These responses were found to be temporal (lasting 3 days). The post-challenge primed state was examined using A. thaliana challenged with a root pathogen. Similarly, defence response-related genes, such as PR1 and NPR1, were up-regulated and ROS production was activated (lasting 5 days). This study found that SWE induces plant priming-like responses by (i) up-regulating genes associated with plant defence responses and (ii) increasing production of ROS associated with signalling responses.

Phaeophyceaean (Brown Algal) Extracts Activate Plant Defense Systems in Arabidopsis thaliana Challenged With Phytophthora cinnamomi.

Seaweed extracts are important sources of plant biostimulants that boost agricultural productivity to meet current world demand. The ability of seaweed extracts based on either of the Phaeophyceaean species Ascophyllum nodosum or Durvillaea potatorum to enhance plant growth or suppress plant disease have recently been shown. However, very limited information is available on the mechanisms of suppression of plant disease by such extracts. In addition, there is no information on the ability of a combination of extracts from A. nodosum and D. potatorum to suppress a plant pathogen or to induce plant defense. The present study has explored the transcriptome, using RNA-seq, of Arabidopsis thaliana following treatment with extracts from the two species, or a mixture of both, prior to inoculation with the root pathogen Phytophthora cinnamomi. Following inoculation, five time points (0-24 h post-inoculation) that represented early stages in the interaction of the pathogen with its host were assessed for each treatment and compared with their respective water controls. Wide scale transcriptome reprogramming occurred predominantly related to phytohormone biosynthesis and signaling, changes in metabolic processes and cell wall biosynthesis, there was a broad induction of proteolysis pathways, a respiratory burst and numerous defense-related responses were induced. The induction by each seaweed extract of defense-related genes coincident with the time of inoculation showed that the plants were primed for defense prior to infection. Each seaweed extract acted differently in inducing plant defense-related genes. However, major systemic acquired resistance (SAR)-related genes as well as salicylic acid-regulated marker genes (PR1, PR5, and NPR1) and auxin associated genes were found to be commonly up-regulated compared with the controls following treatment with each seaweed extract. Moreover, each seaweed extract suppressed P. cinnamomi growth within the roots of inoculated A. thaliana by the early induction of defense pathways and likely through ROS-based signaling pathways that were linked to production of ROS. Collectively, the RNA-seq transcriptome analysis revealed the induction by seaweed extracts of suites of genes that are associated with direct or indirect plant defense in addition to responses that require cellular energy to maintain plant growth during biotic stress.

Increased growth response of strawberry roots to a commercial extract from Durvillaea potatorum and Ascophyllum nodosum.

The withdrawal of soil fumigants like methyl bromide is forcing strawberry growers to consider supplementary and alternative ways of producing crops. In addition to controlling soil-borne pests, soil fumigation causes an increased growth response in strawberry roots, and the use of biostimulants may offer an alternative to replace this response. We tested the hypothesis that treatment with a commercial extract (Seasol®) from the seaweeds Duvillaea potatorum and Ascophyllum nodosum can increase root growth, and transplant (runner) and fruit yields of strawberry. From 2014 to 2016, we conducted three field trials on strawberry farms in the nursery sector at Toolangi and in the fruiting sector at Coldstream in Victoria, Australia. We applied the seaweed extract as a monthly drench (10 L ha-1) to two cultivars of strawberry ('Albion' and 'Fortuna'), compared with an untreated control. In the nursery sector, use of the extract significantly increased the density of secondary roots (feeder roots) on harvested runners by up to 22%. Treatment with the extract also significantly increased yields of marketable runners by 8-19%. In the fruit sector, use of the extract significantly increased the root length density (root length per volume of soil) of strawberry plants by 38% and marketable fruit yields by 8%. Root length density at final harvest and marketable fruit yield of strawberry were highly correlated (r = 0.94). This relationship provides an insight into the mode of action of seaweed extracts and is discussed. Overall, the results show the potential benefits of the integrated use of seaweed extracts in strawberry production across the nursery and fruit sectors, and their promise for supplementing or replacing the increased growth response provided by soil fumigants.

A temperature-sensitive allele of a putative mRNA splicing helicase down-regulates many cell wall genes and causes radial swelling in Arabidopsis thaliana.

The putative RNA helicase encoded by the Arabidopsis gene At1g32490 is a homolog of the yeast splicing RNA helicases Prp2 and Prp22. We isolated a temperature-sensitive allele (rsw12) of the gene in a screen for root radial swelling mutants. Plants containing this allele grown at the restrictive temperature showed weak radial swelling, were stunted with reduced root elongation, and contained reduced levels of cellulose. The role of the protein was further explored by microarray analysis. By using both fold change cutoffs and a weighted gene coexpression network analysis (WGCNA) to investigate coexpression of genes, we found that the radial swelling phenotype was not linked to genes usually associated with primary cell wall biosynthesis. Instead, the mutation has strong effects on expression of secondary cell wall related genes. Many genes potentially associated with secondary walls were present in the most significant WGCNA module, as were genes coding for arabinogalactans and proteins with GPI anchors. The proportion of up-regulated genes that possess introns in rsw12 was above that expected if splicing was unrelated to the activity of the RNA helicase, suggesting that the helicase does indeed play a role in splicing in Arabidopsis. The phenotype may be due to a change in the expression of one or more genes coding for cell wall proteins.

Applications of seaweed extracts in Australian agriculture: past, present and future.

A rapidly growing world population has highlighted the need to significantly increase food production in the context of a world with accelerating soil and water shortages as well as climatic stressors. This situation has generated new interest in the application of liquid seaweed extracts because of their potent plant growth-enhancing properties through metabolic benefits, triggering disease response pathways and increasing stress tolerance. The basis for these benefits is complex and poorly understood. Liquid seaweed extracts are complex and have been demonstrated to possess novel mechanisms for increasing crop productivity. The benefits of seaweed extracts to crops have previously been reviewed in the context of the northern hemisphere, but not in the context of Australia, its crops and unique stressors. This review considers the application of seaweed extracts in Australian agriculture by (i) introducing the history of the Australian liquid seaweed extract industry and (ii) focusing on evidence of Australian research related to seaweed extract composition, plant growth properties during plant establishment, pathogenic disease and new approaches to phenotyping the biological efficacy of seaweed extracts. This type of research is essential for future Australian agriculture to develop effective strategies for the use of liquid seaweed extracts.

A novel isoform of sucrose synthase is targeted to the cell wall during secondary cell wall synthesis in cotton fiber.

Sucrose (Suc) synthase (Sus) is the major enzyme of Suc breakdown for cellulose biosynthesis in cotton (Gossypium hirsutum) fiber, an important source of fiber for the textile industry. This study examines the tissue-specific expression, relative abundance, and temporal expression of various Sus transcripts and proteins present in cotton. A novel isoform of Sus (SusC) is identified that is expressed at high levels during secondary cell wall synthesis in fiber and is present in the cell wall fraction. The phylogenetic relationships of the deduced amino acid sequences indicate two ancestral groups of Sus proteins predating the divergence of monocots and dicots and that SusC sequences form a distinct branch in the phylogeny within the dicot-specific clade. The subcellular location of the Sus isoforms is determined, and it is proposed that cell wall-localized SusC may provide UDP-glucose for cellulose and callose synthesis from extracellular sugars.

A CESA from Griffithsia monilis (Rhodophyta, Florideophyceae) has a family 48 carbohydrate-binding module.

Cellulose synthases form rosette terminal complexes in the plasma membranes of Streptophyta and various linear terminal complexes in other taxa. The sequence of a putative CESA from Griffithsia monilis (Rhodophyta, Floridiophyceae) was deduced using a cloning strategy involving degenerate primers, a cDNA library screen, and 5' and 3' rapid amplification of cDNA ends (RACE). RACE identified two alternative transcriptional starts and four alternative polyadenylation sites. The first translation start codon provided an open reading frame of 2610 bp encoding 870 amino acids and was PCR amplified without introns from genomic DNA. Southern hybridization indicated one strongly hybridizing gene with possible weakly related genes or pseudogenes. Amino acid sequence analysis identified a family 48 carbohydrate-binding module (CBM) upstream of the protein's first predicted transmembrane domain. There are broad similarities in predicted 3D structures of the family 48 modules from CESA, from several glycogen- and starch-binding enzymes, and from protein kinases, but there are substitutions at some residues thought to be involved in ligand binding. The module in G. monilis CESA will be on the cytoplasmic face of the plasma membrane so that it could potentially bind either low molecular weight ligands or starch which is cytosolic rather than inside membrane-bound plastids in red algae. Possible reasons why red algal CESAs have evolved family 48 modules perhaps as part of a system to regulate cellulose synthase activity in relation to cellular carbohydrate status are briefly discussed.

Meta-analysis of cotton fiber quality QTLs across diverse environments in a Gossypium hirsutum x G. barbadense RIL population.

BACKGROUND: Cotton fibers (produced by Gossypium species) are the premier natural fibers for textile production. The two tetraploid species, G. barbadense (Gb) and G. hirsutum (Gh), differ significantly in their fiber properties, the former having much longer, finer and stronger fibers that are highly prized. A better understanding of the genetics and underlying biological causes of these differences will aid further improvement of cotton quality through breeding and biotechnology. We evaluated an inter-specific Gh x Gb recombinant inbred line (RIL) population for fiber characteristics in 11 independent experiments under field and glasshouse conditions. Sites were located on 4 continents and 5 countries and some locations were analyzed over multiple years. RESULTS: The RIL population displayed a large variability for all major fiber traits. QTL analyses were performed on a per-site basis by composite interval mapping. Among the 651 putative QTLs (LOD > 2), 167 had a LOD exceeding permutation based thresholds. Coincidence in QTL location across data sets was assessed for the fiber trait categories strength, elongation, length, length uniformity, fineness/maturity, and color. A meta-analysis of more than a thousand putative QTLs was conducted with MetaQTL software to integrate QTL data from the RIL and 3 backcross populations (from the same parents) and to compare them with the literature. Although the global level of congruence across experiments and populations was generally moderate, the QTL clustering was possible for 30 trait x chromosome combinations (5 traits in 19 different chromosomes) where an effective co-localization of unidirectional (similar sign of additivity) QTLs from at least 5 different data sets was observed. Most consistent meta-clusters were identified for fiber color on chromosomes c6, c8 and c25, fineness on c15, and fiber length on c3. CONCLUSIONS: Meta-analysis provided a reliable means of integrating phenotypic and genetic mapping data across multiple populations and environments for complex fiber traits. The consistent chromosomal regions contributing to fiber quality traits constitute good candidates for the further dissection of the genetic and genomic factors underlying important fiber characteristics, and for marker-assisted selection.

Transcript profiling during fiber development identifies pathways in secondary metabolism and cell wall structure that may contribute to cotton fiber quality.

A global gene expression profiling study at different stages of fiber development was undertaken on two cotton species cultivated for fiber, Gossypium hirsutum (L.) and G. barbadense (L.). A large proportion of the genome was expressed during both fiber elongation and subsequent secondary cell wall thickening. There was a major shift in abundance of transcripts for gene regulation, cell organization and metabolism between fiber elongation and fiber thickening that was fundamentally similar in both species. Each stage had its own distinctive features represented by specific metabolic and regulatory genes, a number of which have been noted previously. Many of the genes expressed in the fibers were of a similar type and developmental expression to those seen in other fiber-producing plants, indicating a conservation of mechanisms of cell elongation and wall thickening across diverse plant genera. Secondary metabolism and pectin synthesis and modification genes were amongst the most statistically significant differentially expressed categories between the two species during fiber elongation. The gene profiles of the fiber thickening stage, however, were almost identical between the two species, suggesting that their different final fiber quality properties may be established at earlier stages of fiber development. Expression levels of representative phenylpropanoid and pectin modification genes showed high correlations with specific fiber properties in an inter-specific cotton recombinant inbred line (RIL) population, supporting a role in determining fiber quality.

Arabidopsis dynamin-like protein DRP1A: a null mutant with widespread defects in endocytosis, cellulose synthesis, cytokinesis, and cell expansion.

Dynamin-related proteins are large GTPases that deform and cause fission of membranes. The DRP1 family of Arabidopsis thaliana has five members of which DRP1A, DRP1C, and DRP1E are widely expressed. Likely functions of DRP1A were identified by studying rsw9, a null mutant of the Columbia ecotype that grows continuously but with altered morphology. Mutant roots and hypocotyls are short and swollen, features plausibly originating in their cellulose-deficient walls. The reduction in cellulose is specific since non-cellulosic polysaccharides in rsw9 have more arabinose, xylose, and galactose than those in wild type. Cell plates in rsw9 roots lack DRP1A but still retain DRP1E. Abnormally placed and often incomplete cell walls are preceded by abnormally curved cell plates. Notwithstanding these division abnormalities, roots and stems add new cells at wild-type rates and organ elongation slows because rsw9 cells do not grow as long as wild-type cells. Absence of DRP1A reduces endocytotic uptake of FM4-64 into the cytoplasm of root cells and the hypersensitivity of elongation and radial swelling in rsw9 to the trafficking inhibitor monensin suggests that impaired endocytosis may contribute to the development of shorter fatter roots, probably by reducing cellulose synthesis.

A mutation in an Arabidopsis ribose 5-phosphate isomerase reduces cellulose synthesis and is rescued by exogenous uridine.

The Arabidopsis radial swelling mutant rsw10 showed ballooning of root trichoblasts, a lower than wild-type level of cellulose and altered levels of some monosaccharides in non-cellulosic polysaccharides. Map-based cloning showed that the mutated gene (At1g71100) encodes a ribose 5-phosphate isomerase (RPI) and that the rsw10 mutation replaces a conserved glutamic acid residue with lysine. Although RPI is intimately involved with many biochemical pathways, media supplementation experiments suggest that the visible phenotype results from a defect in the production of pyrimidine-based sugar-nucleotide compounds, most likely uridine 5'-diphosphate-glucose, the presumed substrate of cellulose synthase. Two of three RPI sequences in the nuclear genome are cytoplasmic, while the third has a putative chloroplast transit sequence. The sequence encoding both cytoplasmic enzymes could complement the mutation when expressed behind the CaMV 35S promoter, while fusion of the RSW10 promoter region to the GUS reporter gene established that the gene is expressed in many aerial tissues as well as the roots. The prominence of the rsw10 phenotype in roots probably reflects RSW10 being the only cytosolic RPI in this tissue and the gene encoding the plastid RPI being relatively weakly expressed. We could not, however, detect a decrease in total RPI activity in root extracts. The rsw10 phenotype is prominent near the root tip where cells undergo division, endoreduplication and cell expansion and so are susceptible to a restriction in de novo pyrimidine production. The two cytosolic RPIs probably arose in an ancient duplication event, their present expression patterns representing subfunctionalization of the expression of the original ancestral gene.

The cellulose-deficient Arabidopsis mutant rsw3 is defective in a gene encoding a putative glucosidase II, an enzyme processing N-glycans during ER quality control.

rsw3 is a temperature-sensitive mutant of Arabidopsis thaliana showing radially swollen roots and a deficiency in cellulose. The rsw3 gene was identified by a map-based strategy, and shows high similarity to the catalytic alpha-subunits of glucosidase II from mouse, yeast and potato. These enzymes process N-linked glycans in the ER, so that they bind and then release chaperones as part of the quality control pathway, ensuring correct protein folding. Putative beta-subunits for the glucosidase II holoenzyme identified in the Arabidopsis and rice genomes share characteristic motifs (including an HDEL ER-retention signal) with beta-subunits in mammals and yeast. The genes encoding the putative alpha- and beta-subunits are single copy and, like the rsw3 phenotype, widely expressed. rsw3 reduces cell number more strongly than cell size in stamen filaments and probably stems. Most features of the rsw3 phenotype are shared with other cellulose-deficient mutants, but some--notably, production of multiple rosettes and a lack of secreted seed mucilage--are not and may reflect glucosidase II affecting processes other than cellulose synthesis. The rsw3 root phenotype develops more slowly than the rsw1 and rsw2 phenotypes when seedlings are transferred to the restrictive temperature. This is consistent with rsw3 reducing glycoprotein delivery from the ER to the plasma membrane whereas rsw1 and rsw2 act more rapidly by affecting the properties of already delivered enzymes.