Publications - Stress and Develop Biology

IntroductionHistorical OutlineChemical StructuresNomenclature and Structure of MetallothioneinsPhytochelatins and Phytochelatin–Metal ComplexesStructural Properties of MetallochaperonesChemical Analysis and DetectionMetallothioneinsPhytochelatinsOccurrenceMetallothioneinsPhytochelatinsMetallochaperonesFunctionsMetal Homeostasis and the Role of MetallochaperonesBuffering and DetoxificationPhytochelatin FunctionsMetallothionein FunctionsPhysiologyMetallothionein Localization and IsoformsLocalization and Compartmentation of Phytochelatin SynthesisBiochemistryMetal‐binding Characteristics of MetallothioneinsBiochemistry of Phytochelatin SynthesisMolecular GeneticsMetallothionein Genes and Their RegulationPhytochelatin Synthase GenesBiotechnological ApplicationsPatentsOutlook and Perspectives

0

0

The HrpZ gene product, harpin, is an export substrate of the type III secretion system of phytopathogenic Pseudomonas syringae. The role of this protein in pathogenesis is largely unknown. We previously determined that HrpZ binds to lipids and can form cation pores in synthetic lipid bilayers. Such pore-forming activity may allow nutrient release during bacterial colonisation of host plants. In addition. HrpZ is known to trigger plant defence responses in a variety of plants, such as tobacco. We have previously also characterised a binding site in tobacco plasma membranes that likely mediates HrpZ-induced defence responses. In order to reconcile these findings, we pose the question as to whether the activation of plant defence responses by HrpZ is mediated through a “classical” receptor perception mode or if plant membrane perturbation through the inherent pore-forming activity of HrpZ may induce defence responses. As defence in parsley cells can be induced both in a receptor-mediated manner or through ionophores these cells served as an ideal system for our analysis. We first performed ligand binding studies to characterise the presence of a binding site/receptor. We further digested HrpZ with endopeptidases and used subfragments of HrpZ to assess the elicitor-active domain of HrpZ. A C-terminal region of HrpZ appears to be sufficient to elicit plant defence responses. A novel assay involving dye-loaded liposomes was developed to validate previous electrophysiological findings on HrpZ-mediated cation pore formation. More importantly, this assay was used to establish if the elicitor-active C-terminal fragment of HrpZ could form pores. Our findings suggest that the structural requirements for ion pore formation and activation of plant defence responses by HrpZ are different. Thus, ion pore formation alone may not explain the activation of plant defence by HrpZ.

0

0

0

0

The Arabidopsis thaliana cDNA, KAT1 encodes a hyperpolarization-activated K+ (K+ in ) channel. In the present study, we identify and characterize dominant negative point mutations that suppress K+ in channel function. Effects of two mutations located in the H5 region of KAT1, at positions 256 (T256R) and 262 (G262K), were studied. The co-expression of either T256R or G262K mutants with KAT1 produced an inhibition of K+ currents upon membrane hyperpolarization. The magnitude of this inhibition was dependent upon the molar ratio of cRNA for wild-type to mutant channel subunits injected. Inhibition of KAT1 currents by the co-expression of T256R or G262K did not greatly affect the ion selectivity of residual currents for Rb+, Na+, Li+, or Cs+. When T256R or G262K were co-expressed with a different K+ channel, AKT2, an inhibition of the channel currents was also observed. Voltage-dependent Cs+ block experiments with co-expressed wild type, KAT1 and AKT2, channels further indicated that KAT1 and AKT2 formed heteromultimers. These data show that AKT2 and KAT1 are able to co-assemble and suggest that suppression of channel function can be pursued in vivo by the expression of the dominant negative K + in channel mutants described here.