Vascular wilt caused by Fusarium oxysporum f. sp. ciceri Race 1 (Foc1) is a serious disease of chickpea (Cicer arietinum L.) accounting for approximately 10-15% annual crop loss. The fungus invades the plant via roots, colonizes the xylem vessels and prevents the upward translocation of water and nutrients, finally resulting in wilting of the entire plant. Although comparative transcriptomic profiling have highlighted some important signaling molecules, but proteomic studies involving chickpea-Foc1 are limited. The present study focuses on comparative root proteomics of susceptible (JG62) and resistant (WR315) chickpea genotypes infected with Foc1, to understand the mechanistic basis of susceptibility and/or resistance.
The differential and unique proteins of both genotypes were identified at 48h, 72h, and 96h post Foc1 inoculation. 2D PAGE analyses followed by MALDI-TOF MS and MS/MS identified 100 differentially (>1.5fold<, p < 0.05) or uniquely expressed proteins. These proteins were further categorized into 10 functional classes and grouped into GO (gene ontology) categories. Network analyses of identified proteins revealed intra and inter relationship of these proteins with their neighbors as well as their association with different defense signaling pathways. qRT-PCR analyses were performed to correlate the mRNA and protein levels of some proteins of representative classes.
The differential and unique proteins identified indicate their involvement in early defense signaling of the host. Comparative analyses of expression profiles of obtained proteins suggest that albeit some common components participate in early defense signaling in both susceptible and resistant genotypes, but their roles and regulation differ in case of compatible and/or incompatible interactions. Thus, functional characterization of identified PR proteins (PR1, BGL2, TLP), Trypsin protease inhibitor, ABA responsive protein, cysteine protease, protein disulphide isomerase, ripening related protein and albumins are expected to serve as important molecular components for biotechnological application and development of sustainable resistance against Foc1.
Necrosis and ethylene-inducing peptide 1 (Nep1)-like proteins (NLPs) are secreted by a wide range of plant-associated microorganisms. They are best known for their cytotoxicity in dicot plants that leads to the induction of rapid tissue necrosis and plant immune responses. The biotrophic downy mildew pathogen Hyaloperonospora arabidopsidis encodes 10 different noncytotoxic NLPs (HaNLPs) that do not cause necrosis. We discovered that these noncytotoxic NLPs, however, act as potent activators of the plant immune system in Arabidopsis thaliana. Ectopic expression of HaNLP3 in Arabidopsis triggered resistance to H. arabidopsidis, activated the expression of a large set of defense-related genes, and caused a reduction of plant growth that is typically associated with strongly enhanced immunity. N- and C-terminal deletions of HaNLP3, as well as amino acid substitutions, pinpointed to a small central region of the protein that is required to trigger immunity, indicating the protein acts as a microbe-associated molecular pattern (MAMP). This was confirmed in experiments with a synthetic peptide of 24 aa, derived from the central part of HaNLP3 and corresponding to a conserved region in type 1 NLPs that induces ethylene production, a well-known MAMP response. Strikingly, corresponding 24-aa peptides of fungal and bacterial type 1 NLPs were also able to trigger immunity in Arabidopsis. The widespread phylogenetic distribution of type 1 NLPs makes this protein family (to our knowledge) the first proteinaceous MAMP identified in three different kingdoms of life.
In this review, various molecular tools used in filamentous fungi are compared and discussed, including methods for genetic transformation (e.g., protoplast transformation, electroporation, and microinjection), the construction of random mutant libraries (e.g., restriction enzyme mediated integration, transposon arrayed gene knockout, and Agrobacterium tumefaciens mediated transformation), and the analysis of gene function (e.g., RNA interference and transcription activator-like effector nucleases). We also focused on practical strategies that could enhance the efficiency of genetic manipulation in filamentous fungi, such as choosing a proper screening system and marker genes, assembling target-cassettes or vectors effectively, and transforming into strains that are deficient in the nonhomologous end joining pathway.
ntricate mechanisms discriminate between friends and foes in plants. Plant organs deploy overlapping and distinct protection strategies. Despite vulnerability to a plethora of pathogens, the growing tips of plants grow bacteria free. The shoot apical meristem (SAM) is among three stem cells niches, a self-renewable reservoir for the future organogenesis of leaf, stem, and flowers. How plants safeguard this high value growth target from infections was not known until now. Recent reports find the stem cell secreted 12-amino acid peptide CLV3p (CLAVATA3 peptide) is perceived by FLS2 (FLAGELLIN SENSING 2) receptor and activates the transcription of immunity and defense marker genes. No infection in the SAM of wild type plants and bacterial infection in clv3 and fls2 mutants illustrate this natural protection against infections. Cytokinins (CKs) are enriched in the SAM and regulate meristem activities by their involvement in stem cell signaling networks. Auxin mediates plant susceptibility to pathogen infections while CKs boost plant immunity. Here, in addition to the stem-cell-triggered immunity we also highlight a potential link between CK signaling and CLV3p mediated immune response in the SAM.
Highlights•The current knowledge about substrates of the key stress regulatory kinase MPK3 is compiled.•Experimental approaches in mitogen-activated protein kinase (MAPK) research, and their pro and cons are described.•General patterns of the occurrence and distribution of peptide motifs related to MAPK control are defined.•Models of how the MAPK recognises, binds and phosphorylates its targets are proposed.
Mitogen-activated protein kinase (MAPK) cascades are universal, evolutionary conserved signalling modules, which translate environmental information into appropriate responses via phosphorylation of their substrate proteins. In Arabidopsis, the MAPK MPK3 regulates numerous cellular processes, including the adaptation to abiotic and biotic stresses. The molecular steps immediately downstream of MPK3 induction have, therefore, received abundant attention, and a respectable number of MPK3 targets are known by now. These proteins illustrate the substrate promiscuity of MPK3. They also are evidence for how manifold phosphorylation-regulated functions can be. This review presents the current knowledge about the function and regulation of MPK3-targeted proteins, takes a close look at their primary protein sequences, and proposes a model of how MPK3 recognises, binds, and phosphorylates its targets.
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