Employing Genomic Tools to Explore the Molecular Mechanisms behind the Enhancement of Plant Growth and Stress Resilience Facilitated by a Burkholderia Rhizobacterial Strain.

The rhizobacterial strain BJ3 showed 16S rDNA sequence similarity to species within the Burkholderia genus. Its complete genome sequence revealed a 97% match with Burkholderia contaminans and uncovered gene clusters essential for plant-growth-promoting traits (PGPTs). These clusters include genes responsible for producing indole acetic acid (IAA), osmolytes, non-ribosomal peptides (NRPS), volatile organic compounds (VOCs), siderophores, lipopolysaccharides, hydrolytic enzymes, and spermidine. Additionally, the genome contains genes for nitrogen fixation and phosphate solubilization, as well as a gene encoding 1-aminocyclopropane-1-carboxylate (ACC) deaminase. The treatment with BJ3 enhanced root architecture, boosted vegetative growth, and accelerated early flowering in Arabidopsis. Treated seedlings also showed increased lignin production and antioxidant capabilities, as well as notably increased tolerance to water deficit and high salinity. An RNA-seq transcriptome analysis indicated that BJ3 treatment significantly activated genes related to immunity induction, hormone signaling, and vegetative growth. It specifically activated genes involved in the production of auxin, ethylene, and salicylic acid (SA), as well as genes involved in the synthesis of defense compounds like glucosinolates, camalexin, and terpenoids. The expression of AP2/ERF transcription factors was markedly increased. These findings highlight BJ3's potential to produce various bioactive metabolites and its ability to activate auxin, ethylene, and SA signaling in Arabidopsis, positioning it as a new Burkholderia strain that could significantly improve plant growth, stress resilience, and immune function.

Examining the Transcriptomic and Biochemical Signatures of Bacillus subtilis Strains: Impacts on Plant Growth and Abiotic Stress Tolerance.

Rhizobacteria from various ecological niches display variations in physiological characteristics. This study investigates the transcriptome profiling of two Bacillus subtilis strains, BsCP1 and BsPG1, each isolated from distinct environments. Gene expression linked to the synthesis of seven types of antibiotic compounds was detected in both BsCP1 and BsPG1 cultures. Among these, the genes associated with plipastatin synthesis were predominantly expressed in both bacterial strains. However, genes responsible for the synthesis of polyketide, subtilosin, and surfactin showed distinct transcriptional patterns. Additionally, genes involved in producing exopolysaccharides (EPS) showed higher expression levels in BsPG1 than in BsCP1. Consistently with this, a greater quantity of EPS was found in the BsPG1 culture compared to BsCP1. Both bacterial strains exhibited similar effects on Arabidopsis seedlings, promoting root branching and increasing seedling fresh weight. However, BsPG1 was a more potent enhancer of drought, heat, and copper stress tolerance than BsCP1. Treatment with BsPG1 had a greater impact on improving survival rates, increasing starch accumulation, and stabilizing chlorophyll content during the post-stress stage. qPCR analysis was used to measure transcriptional changes in Arabidopsis seedlings in response to BsCP1 and BsPG1 treatment. The results show that both bacterial strains had a similar impact on the expression of genes involved in the salicylic acid (SA) and jasmonic acid (JA) signaling pathways. Likewise, genes associated with stress response, root development, and disease resistance showed comparable responses to both bacterial strains. However, treatment with BsCP1 and BsPG1 induced distinct activation of genes associated with the ABA signaling pathway. The results of this study demonstrate that bacterial strains from different ecological environments have varying abilities to produce beneficial metabolites for plant growth. Apart from the SA and JA signaling pathways, ABA signaling triggered by PGPR bacterial strains could play a crucial role in building an effective resistance to various abiotic stresses in the plants they colonize.

Exploring the Biologically Active Metabolites Produced by Bacillus cereus for Plant Growth Promotion, Heat Stress Tolerance, and Resistance to Bacterial Soft Rot in Arabidopsis.

Eight gene clusters responsible for synthesizing bioactive metabolites associated with plant growth promotion were identified in the Bacillus cereus strain D1 (BcD1) genome using the de novo whole-genome assembly method. The two largest gene clusters were responsible for synthesizing volatile organic compounds (VOCs) and encoding extracellular serine proteases. The treatment with BcD1 resulted in an increase in leaf chlorophyll content, plant size, and fresh weight in Arabidopsis seedlings. The BcD1-treated seedlings also accumulated higher levels of lignin and secondary metabolites including glucosinolates, triterpenoids, flavonoids, and phenolic compounds. Antioxidant enzyme activity and DPPH radical scavenging activity were also found to be higher in the treated seedlings as compared with the control. Seedlings pretreated with BcD1 exhibited increased tolerance to heat stress and reduced disease incidence of bacterial soft rot. RNA-seq analysis showed that BcD1 treatment activated Arabidopsis genes for diverse metabolite synthesis, including lignin and glucosinolates, and pathogenesis-related proteins such as serine protease inhibitors and defensin/PDF family proteins. The genes responsible for synthesizing indole acetic acid (IAA), abscisic acid (ABA), and jasmonic acid (JA) were expressed at higher levels, along with WRKY transcription factors involved in stress regulation and MYB54 for secondary cell wall synthesis. This study found that BcD1, a rhizobacterium producing VOCs and serine proteases, is capable of triggering the synthesis of diverse secondary metabolites and antioxidant enzymes in plants as a defense strategy against heat stress and pathogen attack.

Rhizobacterial Bacillus mycoides functions in stimulating the antioxidant defence system and multiple phytohormone signalling pathways to regulate plant growth and stress tolerance.

AIMS: To analyse effects and mechanisms of plant growth promotion mediated by Bacillus mycoides strain A3 (BmA3), in Arabidopsis thaliana seedlings. METHODS AND RESULTS: Bacillus mycoides strain A3 (BmA3) isolated from the bamboo rhizosphere produced phytohormones, including indole-3-acetic acid (IAA) and gibberellic acid (GA), and exhibited phosphate solubilization and radical scavenging activities. A. thaliana seedlings inoculated with BmA3 exhibited an altered root architecture including an increased number of lateral roots and root hairs. Likewise, enhanced photosynthetic efficiency through the accumulation of higher levels of chlorophyll and starch, and increased plant size and fresh weight were observed in the BmA3-treated seedlings. This bacterial inoculation stimulated the antioxidant defence system by increasing the activities of catalase (CAT), guaiacol peroxidase (GPX), ascorbate peroxidase (APX) and phenylalanine ammonia-lyase (PAL). Secondary metabolites, including phenolic compounds, flavonoids and glucosinolates, were induced to higher levels in the BmA3-treated plants. Under drought and heat stresses, lower levels of H2 O2 , malondialdehyde (MDA) and electrolyte leakage were noticed in the treated seedlings. Genes involved in the signalling pathway of jasmonic acid (JA) including MYC2 and lipoxygenase 1 (LOX1) and salicylic acid (SA) including SAR DEFICIENT 1 (SARD1) and CAM-BINDING PROTEIN 60-LIKE G (CBP60G), and the antioxidant defence system including Ascorbate peroxidase (AtAPX) and alternative oxidase (AOX) were upregulated in BmA3-treated plants. Moreover, pathogenesis-related protein 1 (PR-1) and PR-2, marker genes for disease resistance, as well as DREB2A and HsFA2, which function in abiotic stress regulation, were also upregulated. CONCLUSIONS: BmA3 was able to activate JA and SA signalling pathways to induce plant growth and abiotic stress tolerance in A. thaliana seedlings. SIGNIFICANCE AND IMPACT OF STUDY: The plant growth promotion and increased stress tolerance induced by BmA3 were the result of the combined effects of microbial metabolites and activated host plant responses, including phytohormone signalling pathways and antioxidant defence systems.

The role of arabidopsis WDR protein in plant growth and defense strategies.

Evidence indicates that the mechanisms controlling photosynthesis efficiency also regulate plant response to biotic and abiotic stress. Light-induced cell death is genetically maintained for the control of innate immunity. In a recent study we showed that the expression of AtWDR26 was induced by light, multiple plant hormones, and abiotic stress; increased AtWDR26 strongly upregulated gene groups related to chloroplast metabolism, disease resistance, and abiotic stress tolerance. Gain- and loss-of-function analyses in transgenic plants demonstrated the involvement of AtWDR26 in signaling pathways; these controls were osmotic as well as salt stress tolerance. More detailed transcriptome evidence suggested that AtWDR26 was a powerful inducer of gene expression associated with chloroplast metabolism. This included the electron transport chain of the photosystem, carbohydrate synthesis, and enzymatic activity involved in photorespiration. Moreover, genes in auxin synthesis (and perception) constituted a significant portion of those that were upregulated. Gene expression involved in disease resistance, control of cell wall flexibility, Zn uptake, and AP2/ERF transcription factors was also be upregulated. We concluded that AtWDR26 is one component in the regulatory network between light-regulated plant growth and the adaptation response to disease resistance and abiotic stress. Auxin signal acts downstream for AtWDR26 regulation and the adaptation response to biotic and abiotic stress: this occurs through modulating cell wall flexibility, Zn homeostasis, and controlling stress-related transcription factors.

Laminarin modulates the chloroplast antioxidant system to enhance abiotic stress tolerance partially through the regulation of the defensin-like gene expression.

Algae wall polysaccharide, laminarin (Lam), has an established role on induction of plant disease resistance. In this study, application of Lam increased Arabidopsis fresh weight and enhanced tolerance to salt and heat stress by stabilizing chloroplast under adverse environment. Transcriptome analysis indicated that, in addition to induced a large number of genes associated with the host defense, genes involved in the regulation of abiotic stress tolerance mostly the heat stress response constituted the largest group of the up-regulated genes. Lam induced expression of IRT1, ZIP8, and copper transporters involved in transport of Fe, Zn, Cu ions associated with the activity of chloroplast antioxidant system. Lam also up-regulated genes involved in the synthesis of terpenoid, a plastidial-derived secondary metabolite with antioxidant activity. Overexpression of a Lam-induced defensin like 202 (DEFL202) resulted in increased chloroplast stability under salt stress and increased plant growth activity after heat stress. Expression of antioxidant enzymes including SOD and ascorbate peroxidase (APX), photosystem PsbA-D1 and ABA-dependent responsive to desiccation 22 (RD22) was induced to higher levels in the transgenic seedlings. In sum, our results suggest that Lam is an potent inducer for induction of chloroplastic antioxidant activity. Lam affect plant abiotic stress tolerance partially through regulation of the DEFL-mediated pathway.

An Arabidopsis WDR protein coordinates cellular networks involved in light, stress response and hormone signals.

The WD-40 repeat (WDR) protein acts as a scaffold for protein interactions in various cellular events. An Arabidopsis WDR protein exhibited sequence similarity with human WDR26, a scaffolding protein implicated in H2O2-induced cell death in neural cells. The AtWDR26 transcript was induced by auxin, abscisic acid (ABA), ethylene (ET), osmostic stress and salinity. The expression of AtWDR26 was regulated by light, and seed germination of the AtWDR26 overexpression (OE) and seedling growth of the T-DNA knock-out (KO) exhibited altered sensitivity to light. Root growth of the OE seedlings increased tolerance to ZnSO4 and NaCl stresses and were hypersensitive to inhibition of osmotic stress. Seedlings of OE and KO altered sensitivities to multiple hormones. Transcriptome analysis of the transgenic plants overexpressing AtWDR26 showed that genes involved in the chloroplast-related metabolism constituted the largest group of the up-regulated genes. AtWDR26 overexpression up-regulated a large number of genes related to defense cellular events including biotic and abiotic stress response. Furthermore, several members of genes functioning in the regulation of Zn homeostasis, and hormone synthesis and perception of auxin and JA were strongly up-regulated in the transgenic plants. Our data provide physiological and transcriptional evidence for AtWDR26 role in hormone, light and abiotic stress cellular events.

Impacts of size and shape of silver nanoparticles on Arabidopsis plant growth and gene expression.

Silver nanoparticles (AgNPs) are widely used as antibacterial nanomaterials; however, the environmental impacts of AgNPs remain uncertain. In this study, Arabidopsis physiological responses and gene expression were investigated after exposure to 3 different morphologies of AgNPs. The triangular (47 ± 7 nm) and spherical (8 ± 2 nm) AgNPs exhibited the lowest and highest degrees of antimicrobial activity, respectively. The AgNP-induced phenotypic alterations in Arabidopsis were correlated with nanoparticle morphology and size, in which the decahedral AgNPs (45 ± 5 nm) induced the highest degree of root growth promotion (RGP); however, the spherical AgNPs exhibited no RGP and induced the highest levels of anthocyanin accumulation in Arabidopsis seedlings. The decahedral and spherical AgNPs induced the lowest and highest levels of Cu/Zn superoxide dismutase (CSD2) accumulation, respectively. Moreover, 3 morphologies of AgNPs induced protein accumulations including cell-division-cycle kinase 2 (CDC2), protochlorophyllide oxidoreductase (POR), and fructose-1,6 bisphosphate aldolase (FBA). Regarding transcription, the AgNPs induced the gene expression of indoleacetic acid protein 8 (IAA8), 9-cis-epoxycarotenoid dioxygenase (NCED3), and dehydration-responsive RD22. Additional studies have shown that AgNPs antagonized the aminocyclopropane-1-carboxylic acid (ACC)-derived inhibition of root elongation in Arabidopsis seedlings, as well as reduced the expression of ACC synthase 7 (ACS7) and ACC oxidase 2 (ACO2), suggesting that AgNPs acted as inhibitors of ethylene (ET) perception and could interfere with ET biosynthesis. In conclusion, AgNPs induce ROS accumulation and root growth promotion in Arabidopsis. AgNPs activate Arabidopsis gene expression involved in cellular events, including cell proliferation, metabolism, and hormone signaling pathways.

Characterization of a plant-transformation-ready large-insert BIBAC library of Arabidopsis and bombardment transformation of a large-insert BIBAC of the library into tobacco.

Plant-transformation-ready, large-insert binary bacterial artificial chromosome (BIBAC) libraries are of significance for functional and network analysis of large genomic regions, gene clusters, large-spanning genes, and complex loci in the post-genome era. Here, we report the characterization of a plant-transformation-ready BIBAC library of the sequenced Arabidopsis genome for which such a library is not available to the public, the transformation of a large-insert BIBAC of the library into tobacco by biolistic bombardment, and the expression analysis of its containing genes in transgenic plants. The BIBAC library was constructed from nuclear DNA partially digested with BamHI in the BIBAC vector pCLD04541. It contains 6144 clones and has a mean insert size of 108 kb, representing 5.2× equivalents of the Arabidopsis genome or a probability of greater than 99% of obtaining at least one positive clone from the library using a single-copy sequence as a probe. The transformation of the large-insert BIBAC and analyses of the transgenic plants showed that not only did transgenic plants have intact BIBAC DNA, but also could the BIBAC be transmitted stably into progenies and its containing genes be expressed actively. These results suggest that the large-insert BIBAC library, combined with the biolistic bombardment transformation method, could provide a useful tool for large-scale functional analysis of the Arabidopsis genome sequence and applications in plant-molecular breeding.

A harpin-induced ethylene-responsive factor regulates plant growth and responses to biotic and abiotic stresses.

Harpin protein induces disease resistance in diverse plant species. Transcriptome study of tomato (Solanum lycopersicum) genome indicated that harpin activated cellular events of transcription regulation, signaling transduction, stress response, membrane transporting, photosynthesis and cell wall biosynthesis. Among the harpin-induced genes, the expression of tomato ethylene-response factor 5 (designated as SlERF5) was induced to the highest level. The amino acid sequence of SlERF5 was closely related to that of CaEREBP-C4, NtERF4 and NsERF4. Overexpression of SlERF5 in Arabidopsis thaliana activated a large number of genes involved in signaling pathways of disease resistance, redox system, abiotic response and protein phosphorylation. Seedlings of transgenic plants exhibited hypersensitive response to exogenous ABA and increased tolerance to salt stress. In summary, the results of this study suggest that SlERF5 is a transcription activator for genes involved in the responses to ABA, biotic and abiotic stress. Thus, harpin may exert its effect on plant growth and defense response through activation of SlERF5 transcription factors.

Transcriptome analysis of cadmium response in Ganoderma lucidum.

Ganoderma species are white-rot fungi widespread throughout the world. In this study, a wild isolate of Ganoderma lucidum was first collected and its tolerance was tested in a medium containing 3.0 mM CdCl(2). The cDNA-amplified fragment length polymorphism method was conducted to analyze the transcription profiling of this Ganoderma species in response to Cd treatment. In total, 12 925 transcript-derived fragments (TDFs) were amplified using 256 primer combinations. Forty-nine differentially expressed TDFs were confirmed by DNA dot-blot analysis. Northern blot analysis was used to verify the transcription levels of 34 Cd-inducible TDFs. Sequence analysis indicated that genes involved in reactive oxygen species generation, synthesis of sulfur-containing metabolites, translation machinery, DNA repair, transporting system, proteolysis pathway, mitochondria function, and cell wall biosynthesis were upregulated by Cd treatment. Our results provide a genome-wide transcriptome profiling of Cd response in Ganoderma species.

Transcriptome analysis of auxin-regulated genes of Arabidopsis thaliana.

Plant hormone auxin elicits diverse responses in plant growth and development. Accumulated data indicate that the ubiquitin-mediated proteolytic pathway plays a crucial role in transducing auxin signaling. To gain more understanding of the molecular mechanisms underlying auxin action, we performed a comparative transcriptome analysis of auxin responsive genes between Arabidopsis Columbia ecotype and the auxin insensitive mutant eta2 by cDNA-AFLP. Using 256 primer combinations, about 5900 transcript-derived fragments (TDFs) were amplified. Sixty-six differentially expressed TDFs were confirmed by DNA dot blot analysis. Sequence analysis indicated that, a large number of genes involved in transcription regulation or RNA metabolism were identified as auxin-regulated genes. Northern blot analyses confirmed transcription levels of 16 auxin-regulated genes. These genes include various forms of transcription regulators, defense related, RING-type ubiquitin ligases, and glycosyl hydrolase. This study demonstrates that auxin exerts its effect in complex transcriptional networks.

Arabidopsis ETA2, an apparent ortholog of the human cullin-interacting protein CAND1, is required for auxin responses mediated by the SCF(TIR1) ubiquitin ligase.

Auxin response in Arabidopsis thaliana requires the SCF(TIR1) ubiquitin ligase. In response to the hormone, SCF(TIR1) targets members of the auxin/indoleacetic acid (Aux/IAA) family of transcriptional regulators for ubiquitin-mediated proteolysis. To identify additional regulators of SCF(TIR1) activity, we conducted a genetic screen to isolate enhancers of the tir1-1 auxin response defect. Here, we report our analysis of the eta2 mutant. Mutations in ETA2 confer several phenotypes consistent with reduced auxin response. ETA2 encodes the Arabidopsis ortholog of human Cullin Associated and Neddylation-Dissociated (CAND1)/TIP120A, a protein recently identified as a cullin-interacting factor. Previous biochemical studies of CAND1 have suggested that it specifically binds to unmodified CUL1 to negatively regulate SCF assembly. By contrast, we find that ETA2 positively regulates SCF(TIR1) because Aux/IAA protein stability is significantly increased in eta2 mutants. Modification of CUL1 by the RUB1/NEDD8 ubiquitin-like protein has been proposed to free CUL1 from CAND1 and promote SCF assembly. We present double mutant analyses of eta2 axr1 plants indicating that liberating CUL1 from ETA2/CAND1 is not the primary role of the RUB modification pathway in the regulation of SCF activity. Our genetic and molecular analysis of SCF(TIR1) function in eta2 mutants provides novel insight into the role of CAND1 in the regulation of SCF ubiquitin-ligase activity.

Arabidopsis SGT1b is required for SCF(TIR1)-mediated auxin response.

The SCF(TIR1) complex is a central regulator of the auxin response pathway in Arabidopsis. This complex functions as a ubiquitin protein ligase that targets members of the auxin/indoleacetic acid (Aux/IAA) family of transcriptional regulators for ubiquitin-mediated degradation in response to auxin. In an attempt to identify additional factors required for SCF(TIR1) activity, we conducted a genetic screen to isolate enhancers of the auxin response defect conferred by the tir1-1 mutation. Here, we report the identification and characterization of the eta3 mutant. The eta3 mutation interacts synergistically with tir1-1 to strongly enhance all aspects of the tir1 mutant phenotype, including auxin inhibition of root growth, lateral root development, hypocotyl elongation at high temperature, and apical dominance. We isolated the ETA3 gene using a map-based cloning strategy and determined that ETA3 encodes SGT1b. SGT1b was identified recently as a factor involved in plant disease resistance signaling, and SGT1 from barley and tobacco extracts was shown to interact with SCF ubiquitin ligases. We conclude that ETA3/SGT1b is required for the SCF(TIR1)-mediated degradation of Aux/IAA proteins.