Quinones are produced and sensed in all kingdoms of life
. Plants are primary producers of quinone
, but the role of quinone as a signalling agent in plants remains largely unknown. One ...well-documented role of quinone is in the induction of haustoria (specialized feeding structures) in plants that parasitize roots, which occurs in the presence of the host-derived quinone compound 2,6-dimethoxy-1,4-benzoquinone (DMBQ)
. However, how parasitic plants sense DMBQ remains unclear, as is whether nonparasitic plants are capable of sensing quinones. Here we use Arabidopsis thaliana and DMBQ as a model plant and quinone to show that DMBQ signalling occurs in Arabidopsis via elevation of cytosolic Ca
concentration. We performed a forward genetic screen in Arabidopsis that isolated DMBQ-unresponsive mutants, which we named cannot respond to DMBQ 1 (card1). The CANNOT RESPOND TO DMBQ 1 (CARD1; At5g49760, also known as HPCA1) gene encodes a leucine-rich-repeat receptor-like kinase that is highly conserved in land plants. In Arabidopsis, DMBQ triggers defence-related gene expression, and card1 mutants show impaired immunity against bacterial pathogens. In Phtheirospermum japonicum (a plant that parasitizes roots), DMBQ initiates Ca
signalling in the root and is important for the development of the haustorium. Furthermore, CARD1 homologues from this parasitic plant complement DMBQ-induced elevation of cytosolic Ca
concentration in the card1 mutant. Our results demonstrate that plants-unlike animals and bacteria-use leucine-rich-repeat receptor-like kinases for quinone signalling. This work provides insights into the role of quinone signalling and CARD1 functions in plants that help us to better understand the signalling pathways used during the formation of the haustorium in parasitic plants and in plant immunity in nonparasitic plants.
Annexins Laohavisit, Anuphon; Davies, Julia M.
New phytologist,
January 2011, Letnik:
189, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Annexins are multifunctional lipid-binding proteins. Plant annexins are expressed throughout the life cycle and are under environmental control. Their association or insertion into membranes may be ...governed by a range of local conditions (Ca²⁺, pH, voltage or lipid identity) and nonclassical sorting motifs. Protein functions include exocytosis, actin binding, peroxidase activity, callose synthase regulation and ion transport. As such, annexins appear capable of linking Ca²⁺, redox and lipid signalling to coordinate development with responses to the biotic and abiotic environment. Significant advances in plant annexin research have been made in the past 2 yr. Here, we review the basis of annexin multifunctionality and suggest how these proteins may operate in the life and death of a plant cell.
Plants sense microbial signatures via activation of pattern recognition receptors (PPRs), which trigger a range of cellular defences. One response is the closure of plasmodesmata,which reduces ...symplastic connectivity and the capacity for direct molecular exchange between host cells.
Plasmodesmal flux is regulated by a variety of environmental cues but the downstream signalling pathways are poorly defined, especially the way in which calcium regulates plasmodesmal closure.
Here, we identify that closure of plasmodesmata in response to bacterial flagellin, but not fungal chitin, is mediated by a plasmodesmal-localized Ca2+-binding protein Calmodulin-like 41 (CML41). CML41 is transcriptionally upregulated by flg22 and facilitates rapid callose deposition at plasmodesmata following flg22 treatment. CML41 acts independently of other defence responses triggered by flg22 perception and reduces bacterial infection.
We propose that CML41 enables Ca2+-signalling specificity during bacterial pathogen attack and is required for a complete defence response against Pseudomonas syringae.
Salinity (NaCl) stress impairs plant growth and inflicts severe crop losses. In roots, increasing extracellular NaCl causes Ca²⁺ influx to elevate cytosolic free Ca²⁺ (Ca²⁺(cyt)) as a second ...messenger for adaptive signaling. Amplification of the signal involves plasma membrane reduced nicotinamide adenine dinucleotide phosphate oxidase activation, with the resultant reactive oxygen species triggering Ca²⁺ influx. The genetic identities of the Ca²⁺-permeable channels involved in generating the Ca²⁺(cyt) signal are unknown. Potential candidates in the model plant Arabidopsis (Arabidopsis thaliana) include annexin1 (AtANN1). Here, luminescent detection of Ca²⁺(cyt) showed that AtANN1 responds to high extracellular NaCl by mediating reactive oxygen species-activated Ca²⁺ influx across the plasma membrane of root epidermal protoplasts. Electrophysiological analysis revealed that root epidermal plasma membrane Ca²⁺ influx currents activated by NaCl are absent from the Atann1 loss-of-function mutant. Both adaptive signaling and salt-responsive production of secondary roots are impaired in the loss-of-function mutant, thus identifying AtANN1 as a key component of root cell adaptation to salinity.
Plant cell growth and stress signaling require Ca²⁺ influx through plasma membrane transport proteins that are regulated by reactive oxygen species. In root cell growth, adaptation to salinity ...stress, and stomatal closure, such proteins operate downstream of the plasma membrane NADPH oxidases that produce extracellular Superoxide anion, a reactive oxygen species that is readily converted to extracellular hydrogen peroxide and hydroxyl radicals, OH˙ In root cells, extracellular OH˙ activates a plasma membrane Ca²⁺ -permeable conductance that permits Ca²⁺ influx. In Arabidopsis thaliana, distribution of this conductance resembles that of annexin1 (ANN1). Annexins are membrane binding proteins that can form Ca²⁺ -permeable conductances in vitro. Here, the Arabidopsis loss-of-function mutant for annexin1 (Atann1) was found to lack the root hair and epidermal OH˙-activated Ca²⁺ -and K⁺ -permeable conductance. This manifests in both impaired root cell growth and ability to elevate root cell cytosolic free Ca²⁺ in response to OH˙. An OH˙-activated Ca²⁺ conductance is reconstituted by recombinant ANN1 in planar lipid bilayers. ANN1 therefore presents as a novel Ca²⁺-permeable transporter providing a molecular link between reactive oxygen species and cytosolic Ca²⁺ in plants.
The protein expression and purification process is an essential initial step for biochemical analysis of a protein of interest. Traditionally, heterologous protein expression systems (such as
, ...yeast, insect cells, and cell-free) are employed for plant protein expression, although a plant expression system is often desirable for plant proteins, to ensure proper post-translational modifications. Here, we describe a method to express and purify the ectodomain of one of the leucine-rich repeat receptor-like kinase called CARD1/HPCA1, from
apoplastic fluid. First, we express His-tagged CARD1 ectodomain in the apoplastic space of
by the Agroinfiltration method. Then, we collect apoplastic fluids from the leaves and purify the His-tagged protein by Ni
-affinity chromatography. In addition to plant-specific post-translational modifications, protein accumulated in the plant apoplastic space, rather than in the cytosolic space, should be kept under an oxidizing environment. Such an environment will help to maintain the property of intrinsic disulfide bonds in the protein of interest. Further, purification from the apoplastic fluids, rather than the total protein extract, will significantly reduce contaminants (for instance RuBisCO) during protein extraction, and simplify downstream processes. We envisage that our system will be useful for expressing various plant proteins, particularly the apoplastic or extracellular regions of membrane proteins.
Plant annexins are ubiquitous, soluble proteins capable of Ca2+-dependent and Ca2+-independent binding to endomembranes and the plasma membrane. Some members of this multigene family are capable of ...binding to F-actin, hydrolysing ATP and GTP, acting as peroxidases or cation channels. These multifunctional proteins are distributed throughout the plant and throughout the life cycle. Their expression and intracellular localization are under developmental and environmental control. The in vitro properties of annexins and their known, dynamic distribution patterns suggest that they could be central regulators or effectors of plant growth and stress signalling. Potentially, they could operate in signalling pathways involving cytosolic free calcium and reactive oxygen species.
Summary
Photosynthetic microbes exhibit light‐dependent electron export across the cell membrane, which can generate electricity in biological photovoltaic (BPV) devices. How electrons are exported ...remains to be determined; the identification of mechanisms would help selection or generation of photosynthetic microbes capable of enhanced electrical output. We show that plasma membrane NADPH oxidase activity is a significant component of light‐dependent generation of electricity by the unicellular green alga Chlamydomonas reinhardtii. NADPH oxidases export electrons across the plasma membrane to form superoxide anion from oxygen. The C. reinhardtii mutant lacking the NADPH oxidase encoded by RBO1 is impaired in both extracellular superoxide anion production and current generation in a BPV device. Complementation with the wild‐type gene restores both capacities, demonstrating the role of the enzyme in electron export. Monitoring light‐dependent extracellular superoxide production with a colorimetric assay is shown to be an effective way of screening for electrogenic potential of candidate algal strains. The results show that algal NADPH oxidases are important for superoxide anion production and open avenues for optimizing the biological component of these devices.
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