Two Liberibacter Proteins Combine to Suppress Critical Innate Immune Defenses in Citrus.

We adopted a systems-based approach to determine the role of two Candidatus Liberibacter asiaticus (CLas) proteins, LasP 235 and Effector 3, in Huanglongbing (HLB) pathogenesis. While a published work suggests the involvement of these CLas proteins HLB pathogenesis, the exact structure-based mechanism of their action has not been elucidated. We conducted the following experiments to determine the structure-based mechanisms of action. First, we immunoprecipitated the interacting citrus protein partners of LasP 235 and Effector 3 from the healthy and CLas-infected Hamlin extracts and identified them by Liquid Chromatography with tandem mass spectrometry (LC-MS/MS). Second, we performed a split green fluorescent protein (GFP) assay in tobacco to validate that the interactions observed in vitro are also retained in planta. The notable in planta citrus targets of LasP 235 and Effector 3 include citrus innate immune proteins. Third, in vitro and in planta studies were performed to show that LasP 235 and Effector 3 interact with and inhibit the functions of multiple citrus proteins belonging to the innate immune pathways. These inhibitory interactions led to a high level of reactive oxygen species, blocking of bactericidal lipid transfer protein (LTP), and induction of premature programed cell death (PCD), all of which are beneficial to CLas lifecycle and HLB pathogenesis. Finally, we performed molecular dynamics simulations to visualize the interactions of LasP 235 and Effector 3, respectively, with LTP and Kunitz protease inhibitor. This led to the design of an LTP mimic, which sequestered and blocked LasP 235 and rescued the bactericidal activity of LTP thereby proving that LasP 235 , indeed, participates in HLB pathogenesis.

Rational design of antimicrobial peptides targeting Gram-negative bacteria.

Membrane-targeting host antimicrobial peptides (AMPs) can kill or inhibit the growth of Gram-negative bacteria. However, the evolution of resistance among microbes poses a substantial barrier to the long-term utility of the host AMPs. Combining experiment and molecular dynamics simulations, we show that terminal carboxyl capping enhances both membrane insertion and antibacterial activity of an AMP called P1. Furthermore, we show that a bacterial strain with evolved resistance to this peptide becomes susceptible to P1 variants with either backbone capping or lysine-to-arginine substitutions. Our results suggest that cocktails of closely related AMPs may be useful in overcoming evolved resistance.

Coping with inclement weather conditions due to high temperature and water deficit in rice: An insight from genetic and biochemical perspectives.

Climatic fluctuations, temperature extremes, and water scarcity are becoming increasingly unpredictable with the passage of time. Such environmental atrocities have been the scourge of agriculture over the ages, bringing with them poor harvests and threat of famine. Rice production, owing to its high-water requirement for cultivation, is highly vulnerable to the threat of changing climate, particularly prolonged drought and high temperature, individually or in combination. Amidst all the abiotic stresses, heat and drought are considered as the most important concurrent stressors, largely affecting rice yield and productivity under the current scenario. Such threats heighten the need for new breeding and cultivation strategies in generating abiotic stress-resilient rice varieties with better yield potential. Responses of rice to these stresses can be categorized at the morphological, physiological and biochemical levels. This review examines the physiological and molecular mechanism, in the form of up regulation of several defense machineries of rice varieties to cope with drought stress (DS), high temperature stress (HTS), and their combination (DS-HTS). Genotypic differences among rice varieties in their tolerance ability have also been addressed. The review also appraises research studies conducted in rice regarding various phenotypic traits, genetic loci and response mechanisms to stress conditions to help craft new breeding strategies for improved tolerance to DS and HTS, singly or in combination. The review also encompasses the gene regulatory networks and transcription factors, and their cross-talks in mediating tolerance to such stresses. Understanding the epigenetic regulation, involving DNA methylation and histone modification during such hostile situations, will also play a crucial role in our comprehensive understanding of combinatorial stress responses. Taken together, this review consolidates current research and available information on promising rice cultivars with desirable traits as well as advocates synergistic and complementary approaches in molecular and systems biology to develop new rice breeds that favorably respond to DS-HTS-induced abiotic stress.

Phytochrome and Phytohormones: Working in Tandem for Plant Growth and Development.

Being sessile organisms, plants need to continually adapt and modulate their rate of growth and development in accordance with the changing environmental conditions, a phenomenon referred to as plasticity. Plasticity in plants is a highly complex process that involves a well-coordinated interaction between different signaling pathways, the spatiotemporal involvement of phytohormones and cues from the environment. Though research studies are being carried out over the years to understand how plants perceive the signals from changing environmental conditions and activate plasticity, such remain a mystery to be resolved. Among all environmental cues, the light seems to be the stand out factor influencing plant growth and development. During the course of evolution, plants have developed well-equipped signaling system that enables regulation of both quantitative and qualitative differences in the amount of perceived light. Light influences essential developmental switches in plants ranging from germination or transition to flowering, photomorphogenesis, as well as switches in response to shade avoidances and architectural changes occurring during phototropism. Abscisic acid (ABA) is controlling seed germination and is regulated by light. Furthermore, circadian clock adds another level of regulation to plant growth by integrating light signals with different hormonal pathways. MYB96 has been identified as a regulator of circadian gating of ABA-mediated responses in plants by binding to the TIMING OF CAB EXPRESSION 1(TOC1) promoter. This review will present a representative regulatory model, highlight the successes achieved in employing novel strategies to dissect the levels of interaction and provide perspective for future research on phytochrome-phytohormones relationships toward facilitating plant growth, development, and function under abiotic-biotic stresses.

Engineering PGPMOs through Gene Editing and Systems Biology: A Solution for Phytoremediation?

In light of extensive urbanization and deforestation, toxic wastes are being released into the atmosphere, causing increased air and soil pollution. Conventional methods of soil remediation are time consuming and labor and cost intensive, rendering them uneconomical to maintain sustainable agriculture. One solution is to use natural resources like plants and microbes for phytoremediation. A thorough systemic knowledge of plant-microbe interactions will allow the use of gene editing and gene manipulation techniques to increase the efficiency of plants in phytoremediation. This Opinion article focuses on gene editing techniques used in plants and microbes for phytoremediation and also emphasizes their effectiveness, advancement, and future implications for sustainable and environmentally friendly agriculture.

Effect of Spectral Quality of Monochromatic LED Lights on the Growth of Artichoke Seedlings.

Indoor farming is becoming a popular alternative approach in food production to meet the demand of a growing world population. Under this production system, artificial light provides the main source of illumination in sustaining plant growth and development. The use of light-emitting diodes (LEDs) is a popular source of artificial light for indoor farms due to its narrow light spectra, modular design and energy efficiency. This study purposely assessed the effect of monochromatic LED light quality on the growth of three varieties of artichoke seedlings compared to greenhouse condition. Spectral quality assessment showed that photosynthetic photon flux density (PPFD) was highest under red LED light, but only a third of the total PPFD under natural light. Seedlings grown under red light showed 60-100% more shoot dry weight and were 67-115% taller than seedlings grown in the greenhouse. However, seedlings under blue or white light conditions showed 67-76% less in biomass compared to greenhouse-grown seedlings. Overall, plant response of seedlings under red light condition was much better compared to greenhouse-grown seedlings emphasizing the importance of red light spectral quality in plant growth and development.

Comparative Metabolome Profile between Tobacco and Soybean Grown under Water-Stressed Conditions.

Understanding how plants respond to water deficit is important in order to develop crops tolerant to drought. In this study, we compare two large metabolomics datasets where we employed a nontargeted metabolomics approach to elucidate metabolic pathways perturbed by progressive dehydration in tobacco and soybean plants. The two datasets were created using the same strategy to create water deficit conditions and an identical metabolomics pipeline. Comparisons between the two datasets therefore reveal common responses between the two species, responses specific to one of the species, responses that occur in both root and leaf tissues, and responses that are specific to one tissue. Stomatal closure is the immediate response of the plant and this did not coincide with accumulation of abscisic acid. A total of 116 and 140 metabolites were observed in tobacco leaves and roots, respectively, while 241 and 207 were observed in soybean leaves and roots, respectively. Accumulation of metabolites is significantly correlated with the extent of dehydration in both species. Among the metabolites that show increases that are restricted to just one plant, 4-hydroxy-2-oxoglutaric acid (KHG) in tobacco roots and coumestrol in soybean roots show the highest tissue-specific accumulation. The comparisons of these two large nontargeted metabolomics datasets provide novel information and suggest that KHG will be a useful marker for drought stress for some members of Solanaceae and coumestrol for some legume species.

A toolbox of genes, proteins, metabolites and promoters for improving drought tolerance in soybean includes the metabolite coumestrol and stomatal development genes.

BACKGROUND: The purpose of this project was to identify metabolites, proteins, genes, and promoters associated with water stress responses in soybean. A number of these may serve as new targets for the biotechnological improvement of drought responses in soybean (Glycine max). RESULTS: We identified metabolites, proteins, and genes that are strongly up or down regulated during rapid water stress following removal from a hydroponics system. 163 metabolites showed significant changes during water stress in roots and 93 in leaves. The largest change was a root-specific 160-fold increase in the coumestan coumestrol making it a potential biomarker for drought and a promising target for improving drought responses. Previous reports suggest that coumestrol stimulates mycorrhizal colonization and under certain conditions mycorrhizal plants have improved drought tolerance. This suggests that coumestrol may be part of a call for help to the rhizobiome during stress. About 3,000 genes were strongly up-regulated by drought and we identified regulators such as ERF, MYB, NAC, bHLH, and WRKY transcription factors, receptor-like kinases, and calcium signaling components as potential targets for soybean improvement as well as the jasmonate and abscisic acid biosynthetic genes JMT, LOX1, and ABA1. Drought stressed soybean leaves show reduced mRNA levels of stomatal development genes including FAMA-like, MUTE-like and SPEECHLESS-like bHLH transcription factors and leaves formed after drought stress had a reduction in stomatal density of 22.34 % and stomatal index of 17.56 %. This suggests that reducing stomatal density may improve drought tolerance. MEME analyses suggest that ABRE (CACGT/CG), CRT/DRE (CCGAC) and a novel GTGCnTGC/G element play roles in transcriptional activation and these could form components of synthetic promoters to drive expression of transgenes. Using transformed hairy roots, we validated the increase in promoter activity of GmWRKY17 and GmWRKY67 during dehydration and after 20 µM ABA treatment. CONCLUSIONS: Our toolbox provides new targets and strategies for improving soybean drought tolerance and includes the coumestan coumestrol, transcription factors that regulate stomatal density, water stress-responsive WRKY gene promoters and a novel DNA element that appears to be enriched in water stress responsive promoters.

The WRKY transcription factor family and senescence in switchgrass.

BACKGROUND: Early aerial senescence in switchgrass (Panicum virgatum) can significantly limit biomass yields. WRKY transcription factors that can regulate senescence could be used to reprogram senescence and enhance biomass yields. METHODS: All potential WRKY genes present in the version 1.0 of the switchgrass genome were identified and curated using manual and bioinformatic methods. Expression profiles of WRKY genes in switchgrass flag leaf RNA-Seq datasets were analyzed using clustering and network analyses tools to identify both WRKY and WRKY-associated gene co-expression networks during leaf development and senescence onset. RESULTS: We identified 240 switchgrass WRKY genes including members of the RW5 and RW6 families of resistance proteins. Weighted gene co-expression network analysis of the flag leaf transcriptomes across development readily separated clusters of co-expressed genes into thirteen modules. A visualization highlighted separation of modules associated with the early and senescence-onset phases of flag leaf growth. The senescence-associated module contained 3000 genes including 23 WRKYs. Putative promoter regions of senescence-associated WRKY genes contained several cis-element-like sequences suggestive of responsiveness to both senescence and stress signaling pathways. A phylogenetic comparison of senescence-associated WRKY genes from switchgrass flag leaf with senescence-associated WRKY genes from other plants revealed notable hotspots in Group I, IIb, and IIe of the phylogenetic tree. CONCLUSIONS: We have identified and named 240 WRKY genes in the switchgrass genome. Twenty three of these genes show elevated mRNA levels during the onset of flag leaf senescence. Eleven of the WRKY genes were found in hotspots of related senescence-associated genes from multiple species and thus represent promising targets for future switchgrass genetic improvement. Overall, individual WRKY gene expression profiles could be readily linked to developmental stages of flag leaves.

Transcriptome profiling of tobacco under water deficit conditions.

Drought is one of the limiting environmental factors that affect crop production. Understanding the molecular basis of how plants respond to this water deficit stress is key to developing drought tolerant crops. In this study we generated time course-based transcriptome profiles of tobacco plants under water deficit conditions using microarray technology. In this paper, we describe in detail the experimental procedures and analyses performed in our study. The data set we generated (available in the NCBI/GEO database under GSE67434) has been analysed to identify genes that are involved in the regulation of tobacco's responses to drought.