Unraveling the involvement of WRKY TFs in regulating plant disease defense signaling.

MAIN CONCLUSION: This review article explores the intricate role, regulation, and signaling mechanisms of WRKY TFs in response to biotic stress, particularly emphasizing their pivotal role in the trophism of plant-pathogen interactions. Transcription factors (TFs) play a vital role in governing both plant defense and development by controlling the expression of various downstream target genes. Early studies have shown the differential expression of certain WRKY transcription factors by microbial infections. Several transcriptome-wide studies later demonstrated that diverse sets of WRKYs are significantly activated in the early stages of viral, bacterial, and fungal infections. Furthermore, functional investigations indicated that overexpression or silencing of certain WRKY genes in plants can drastically alter disease symptoms as well as pathogen multiplication rates. Hence the new aspects of pathogen-triggered WRKY TFs mediated regulation of plant defense can be explored. The already recognized roles of WRKYs include transcriptional regulation of defense-related genes, modulation of hormonal signaling, and participation in signal transduction pathways. Some WRKYs have been shown to directly bind to pathogen effectors, acting as decoys or resistance proteins. Notably, the signaling molecules like salicylic acid, jasmonic acid, and ethylene which are associated with plant defense significantly increase the expression of several WRKYs. Moreover, induction of WRKY genes or heightened WRKY activities is also observed during ISR triggered by the beneficial microbes which protect the plants from subsequent pathogen infection. To understand the contribution of WRKY TFs towards disease resistance and their exact metabolic functions in infected plants, further studies are required. This review article explores the intrinsic transcriptional regulation, signaling mechanisms, and hormonal crosstalk governed by WRKY TFs in plant disease defense response, particularly emphasizing their specific role against different biotrophic, hemibiotrophic, and necrotrophic pathogen infections.

Implications of biosensors and nanobiosensors for the eco-friendly detection of public health and agro-based insecticides: A comprehensive review.

Biosensors, in particular nanobiosensors, have brought a paradigm shift in the detection approaches involved in healthcare, agricultural, and industrial sectors. In accordance with the global expansion in the world population, there has been an increase in the application of specific insecticides for maintaining public health and enhancing agriculture, such as organophosphates, organochlorines, pyrethroids, and carbamates. This has led to the contamination of ground water, besides increasing the chances of biomagnification as most of these insecticides are non-biodegradable. Hence, conventional and more advanced approaches are being devised for the routine monitoring of such insecticides in the environment. This review walks through the implications of biosensors and nanobiosensors, which could offer a wide range of benefits for the detection of the insecticides, quantifying their toxicity status, and versatility in application. Unique eco-friendly nanobiosensors such as microcantilevers, carbon nanotubes, 3D printing organic materials and nylon nano-compounds are some advanced tools that are being employed for the detection of specific insecticides under different conditions. Furthermore, in order to implement a smart agriculture system, nanobiosensors could be integrated into mobile apps and GPS systems for controlling farming in remote areas, which would greatly assist the farmer remotely for crop improvement and maintenance. This review discusses about such tools along with more advanced and eco-friendly approaches that are on the verge of development and could offer a promising alternative for analyte detection in different domains.

MYC2 influences salicylic acid biosynthesis and defense against bacterial pathogens in Arabidopsis thaliana.

Arabidopsis MYC2 is a basic helix-loop-helix transcription factor that works both as a negative and positive regulator of light and multiple hormonal signaling pathways, including jasmonic acid and abscisic acid. Recent studies have suggested the role of MYC2 as a negative regulator of salicylic acid (SA)-mediated defense against bacterial pathogens. By using myc2 mutant and constitutively MYC2-expressing plants, we further show that MYC2 also positively influences SA-mediated defense; whereas, myc2 mutant plants are resistant to virulent pathogens only, MYC2 over-expressing plants are hyper-resistant to multiple virulent and avirulent strains of bacterial pathogens. MYC2 promotes pathogen-induced callose deposition, SA biosynthesis, expression of PR1 gene, and SA-responsiveness. Using bacterially produced MYC2 protein in electrophoretic mobility shift assay (EMSA), we have shown that MYC2 binds to the promoter of several important defense regulators, including PEPR1, MKK4, RIN4, and the second intron of ICS1. MYC2 positively regulates the expression of RIN4, MKK4, and ICS1; however, it negatively regulates the expression of PEPR1. Pathogen inoculation enhances MYC2 association at ICS1 intron and RIN4 promoter. Mutations of MYC2 binding site at ICS1 intron or RIN4 promoter abolish the associated GUS reporter expression. Hyper-resistance of MYC2 over-expressing plants is largely light-dependent, which is in agreement with the role of MYC2 in SA biosynthesis. The results altogether demonstrate that MYC2 possesses dual regulatory roles in SA biosynthesis, SA signaling, pattern-triggered immunity (PTI), and effector-triggered immunity (ETI) in Arabidopsis.

Genome-wide identification and expression analysis of WRKY transcription factors in pearl millet (Pennisetum glaucum) under dehydration and salinity stress.

BACKGROUND: Plants have developed various sophisticated mechanisms to cope up with climate extremes and different stress conditions, especially by involving specific transcription factors (TFs). The members of the WRKY TF family are well known for their role in plant development, phytohormone signaling and developing resistance against biotic or abiotic stresses. In this study, we performed a genome-wide screening to identify and analyze the WRKY TFs in pearl millet (Pennisetum glaucum; PgWRKY), which is one of the most widely grown cereal crops in the semi-arid regions. RESULTS: A total number of 97 putative PgWRKY proteins were identified and classified into three major Groups (I-III) based on the presence of WRKY DNA binding domain and zinc-finger motif structures. Members of Group II have been further subdivided into five subgroups (IIa-IIe) based on the phylogenetic analysis. In-silico analysis of PgWRKYs revealed the presence of various cis-regulatory elements in their promoter region like ABRE, DRE, ERE, EIRE, Dof, AUXRR, G-box, etc., suggesting their probable involvement in growth, development and stress responses of pearl millet. Chromosomal mapping evidenced uneven distribution of identified 97 PgWRKY genes across all the seven chromosomes of pearl millet. Synteny analysis of PgWRKYs established their orthologous and paralogous relationship among the WRKY gene family of Arabidopsis thaliana, Oryza sativa and Setaria italica. Gene ontology (GO) annotation functionally categorized these PgWRKYs under cellular components, molecular functions and biological processes. Further, the differential expression pattern of PgWRKYs was noticed in different tissues (leaf, stem, root) and under both drought and salt stress conditions. The expression pattern of PgWRKY33, PgWRKY62 and PgWRKY65 indicates their probable involvement in both dehydration and salinity stress responses in pearl millet. CONCLUSION: Functional characterization of identified PgWRKYs can be useful in delineating their role behind the natural stress tolerance of pearl millet against harsh environmental conditions. Further, these PgWRKYs can be employed in genome editing for millet crop improvement.

Rice MYC2 (OsMYC2) modulates light-dependent seedling phenotype, disease defence but not ABA signalling.

Arabidopsis MYC2 (AtMYC2) is a bHLH class transcription factor that mediates light-dependent seedling development, disease defence, JA and ABA signalling. AtMYC2 gene modulates hypocotyl elongation and expression of chlorophyll A/B binding protein 1 (CAB1) and rubisco small subunit protein1 (RBCS1) under blue light. The atmyc2 mutants are resistant against virulent bacterial pathogens. MYC2 orthologues from several crop plants have been characterized. The rice gene Os10g42430 has been referred earlier as OsMYC2 and has been shown to promote expression of JA-inducible genes. However, the role of OsMYC2 in seedling development under ABA, dark or light of specific wavelengths was not known. It was also not known whether OsMYC2 complements AtMYC2 function in Arabidopsis. We show here that expression of OsMYC2 in the atmyc2 mutant of Arabidopsis complements the blue-light-mediated defects in hypocotyl elongation and expression of CAB1 and RBCS1. We generated multiple transgenic rice lines for over-expression and RNAi-mediated suppression of OsMYC2. In agreement with AtMYC2 function, OsMYC2 over-expression and RNAi lines showed enhanced and suppressed seedling growth compared to WT plants respectively under blue light, and showed little effect under white light or dark. In agreement with the negative regulatory role of AtMYC2 in disease defence, the RNAi lines showed enhanced resistance against bacterial pathogen Xanthomonas oryzae pv oryzae. However, in contrast to AtMYC2 function, OsMYC2 influences seedling development under red light and show no effect in ABA-mediated seed germination. Thus, the results suggest evolutionarily conserved as well as the distinct role of OsMYC2 in comparison with AtMYC2.

The Arabidopsis thaliana At4g13040 gene, a unique member of the AP2/EREBP family, is a positive regulator for salicylic acid accumulation and basal defense against bacterial pathogens.

The Arabidopsis genome contains a large number of putative transcription factors, containing a DNA binding domain similar to APETALA2/ethylene response element binding protein (AP2/EREBP), for most of which a function is not known. Phylogenetic analysis divides the Apetala 2 (AP2) super-family into 5 major groups: AP2, RAV, ethylene response factor (ERF), dehydration response element binding protein (DREB) and At4g13040. Similar to ERF and DREB, the At4g13040 protein contains only one AP2 domain; however, its structural uniqueness places it into a distinct group. In this article, we report that At4g13040 (referred herein as Apetala 2 family protein involved in SA mediated disease defense 1 - APD1) is an important regulator for SA mediated plant defense. The APD1 gene is upregulated upon pathogen inoculation, exogenous SA application and in the mutant that constitutively activates SA signaling. The T-DNA insertion lines (inserted in the APD1 promoter), which fail to induce expression upon pathogen inoculation, are compromised for resistance against virulent bacterial pathogens and show reduced induction of pathogenesis related 1 gene. Our results suggest that APD1 functions downstream of PAD4 in Arabidopsis and promotes pathogen-induced SA accumulation. Exogenous SA application completely restores the loss-of-resistance phenotype of the apd1 mutant. Thus, APD1 is a positive regulator of disease defense that functions upstream of SA accumulation.

Down-regulation of OsSAG12-1 results in enhanced senescence and pathogen-induced cell death in transgenic rice plants.

Senescence is a highly regulated process accompanied by changes in gene expression. While the mRNA levels of most genes decline, the mRNA levels of specific genes (senescence associated genes, SAGs) increase during senescence. Arabidopsis SAG12 (AtSAG12) gene codes for papain-like cysteine protease. The promoter of AtSAG12 is SA-responsive and reported to be useful to delay senescence by expressing cytokinin biosynthesis gene isopentenyltransferase specifically during senescence in several plants including Arabidopsis, lettuce and rice. The physiological role of AtSAG12 is not known; the homozygous atsag12 mutant neither fails to develop senescenceassociated vacuoles nor shows any morphological phenotype. Through BLAST search using AtSAG12 amino acid sequences as query, we identified a few putative homologues from rice genome (OsSAGs; Oryza sativa SAGs). OsSAG12-1 is the closest homologue of AtSAG12 with 64% similar amino acid composition. Expression of OsSAG12-1 is induced during senescence and pathogen-induced cell death. To evaluate the possible role of OsSAG12-1 we generated RNAi transgenic lines in Japonica rice cultivar TP309. The transgenic lines developed early senescence at varying levels and showed enhanced cell death when inoculated with bacterial pathogen Xanthomonas oryzae pv.oryzae. Our results suggest that OsSAG12-1 is a negative regulator of cell death in rice.

Arabidopsis thaliana FLOWERING LOCUS D is required for systemic acquired resistance.

Localized infection in plants often induces systemic acquired resistance (SAR), which provides long-term protection against subsequent infections. A signal originating in the SAR-inducing organ is transported to the distal organs, where it stimulates salicylic acid (SA) accumulation and priming, a mechanism that results in more robust activation of defenses in response to subsequent pathogen infection. In recent years, several metabolites that promote long-distance SAR signaling have been identified. However, the mechanism or mechanisms by which plants perceive and respond to the SAR signals are largely obscure. Here, we show that, in Arabidopsis thaliana, the FLOWERING LOCUS D (FLD) is required for responding to the SAR signals leading to the systemic accumulation of SA and enhancement of disease resistance. Although the fld mutant was competent in accumulating the SAR-inducing signal, it was unable to respond to the SAR signal that accumulates in petiole exudates of wild-type leaves inoculated with a SAR-inducing pathogen. Supporting FLD's role in systemic SAR signaling, we observed that dehydroabietinal and azelaic acid, two metabolites that, in wild-type plants, promote SAR-associated systemic accumulation of SA and priming, respectively, were unable to promote SAR in the fld mutant. FLD also participates in flowering, where it functions to repress expression of the flowering repressor FLOWERING LOCUS C (FLC). However, epistasis analysis indicates that FLD's function in SAR is independent of FLC.