Hypericum perforatum plant cells reduce Agrobacterium viability during co-cultivation

Plant recalcitrance is the major barrier in developing Agrobacterium-mediated transformation protocols for several important plant species. Despite the substantial knowledge of T-DNA transfer process, very little is known about the factors leading to the plant recalcitrance. Here, we analyzed the basis of Hypericum perforatum L. (HP) recalcitrance to Agrobacterium-mediated transformation using cell suspension culture. When challenged with Agrobacterium, HP cells swiftly produced an intense oxidative burst, a typical reaction of plant defense. Agrobacterium viability started to decline and reached 99% mortality within 12 h, while the plant cells did not suffer apoptotic process. This is the first evidence showing that the reduction of Agrobacterium viability during co-cultivation with recalcitrant plant cells can affect transformation

Overexpression of the Endoplasmic Reticulum Chaperone BiP3 Regulates XA21-Mediated Innate Immunity in Rice

Recognition of pathogen-associated molecular patterns by pattern recognition receptors (PRRs) activates the innate immune response. Although PRR-mediated signaling events are critical to the survival of plants and animals, secretion and localization of PRRs have not yet been clearly elucidated. Here we report the in vivo interaction of the endoplasmic reticulum (ER) chaperone BiP3 with the rice XA21 PRR, which confers resistance to the Gram negative bacterium, Xanthomonas oryzae pv. oryzae (Xoo). We show that XA21 is glycosylated and is primarily localized to the ER and also to the plasma membrane (PM). In BiP3-overexpressing rice plants, XA21-mediated immunity is compromised, XA21 stability is significantly decreased, and XA21 proteolytic cleavage is inhibited. BiP3 overexpression does not affect the general rice defense response, cell death or brassinolide-induced responses. These results indicate that BiP3 regulates XA21 protein stability and processing and that this regulation is critical for resistance to Xoo

Parental Genome Dosage Imbalance Deregulates Imprinting in Arabidopsis

In mammals and in plants, parental genome dosage imbalance deregulates embryo growth and might be involved in reproductive isolation between emerging new species. Increased dosage of maternal genomes represses growth while an increased dosage of paternal genomes has the opposite effect. These observations led to the discovery of imprinted genes, which are expressed by a single parental allele. It was further proposed in the frame of the parental conflict theory that parental genome imbalances are directly mirrored by antagonistic regulations of imprinted genes encoding maternal growth inhibitors and paternal growth enhancers. However these hypotheses were never tested directly. Here, we investigated the effect of parental genome imbalance on the expression of Arabidopsis imprinted genes FERTILIZATION INDEPENDENT SEED2 (FIS2) and FLOWERING WAGENINGEN (FWA) controlled by DNA methylation, and MEDEA (MEA) and PHERES1 (PHE1) controlled by histone methylation. Genome dosage imbalance deregulated the expression of FIS2 and PHE1 in an antagonistic manner. In addition increased dosage of inactive alleles caused a loss of imprinting of FIS2 and MEA. Although FIS2 controls histone methylation, which represses MEA and PHE1 expression, the changes of PHE1 and MEA expression could not be fully accounted for by the corresponding fluctuations of FIS2 expression. Our results show that parental genome dosage imbalance deregulates imprinting using mechanisms, which are independent from known regulators of imprinting. The complexity of the network of regulations between expressed and silenced alleles of imprinted genes activated in response to parental dosage imbalance does not support simple models derived from the parental conflict hypothesis

Inducing Ni Sensitivity in the Ni Hyperaccumulator Plant Alyssum inflatum Nyárády (Brassicaceae) by Transforming with CAX1, a Vacuolar Membrane Calcium Transporter

The importance of calcium in nickel tolerance was studied in the nickel hyperaccumulator plant Alyssum inflatum by gene transformation of CAX1, a vacuolar membrane transporter that reduces cytosolic calcium. CAX1 from Arabidopsis thaliana with a CaMV35S promoter accompanying a kanamycin resistance gene was transferred into A. inflatum using Agrobacterium tumefaciens. Transformed calli were subcultured three times on kanamycin-rich media and transformation was confirmed by PCR using a specific primer for CAX1. At least 10 callus lines were used as a pool of transformed material. Both transformed and untransformed calli were treated with varying concentrations of either calcium (1–15 mM) or nickel (0– 500 lM) to compare their responses to those ions. Increased external calcium generally led to increased callus biomass, however, the increase was greater for untransformed callus. Further, increased external calcium led to increased callus calcium concentrations. Transformed callus was less nickel tolerant than untransformed callus: under increasing nickel concentrations callus relative growth rate was significantly less for transformed callus. Transformed callus also contained significantly less nickel than untransformed callus when exposed to the highest external nickel concentration (200 lM). We suggest that transformation with CAX1 decreased cytosolic calcium and resulted in decreased nickel tolerance. This in turn suggests that, at low cytosolic calcium concentrations, other nickel tolerance mechanisms (e.g., complexation and vacuolar sequestration) are insufficient for nickel tolerance. We propose that high cytosolic calcium is an important mechanism that results in nickel tolerance by nickel hyperaccumulator plants

Characterization of ScMat1, a putative TFIIH subunit from sugarcane

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The general transcription factor TFIIH is a multiprotein complex with different enzymatic activities such as helicase, protein kinase and DNA repair. MAT1 (m,nage A trois 1) is one of the TFIIH subunits that has kinase activity and it is the third subunit of the cyclin-dependent kinase (CDK)-activating kinase (CAK), CDK7- cyclin H. The main objective of this work was to characterize ScMAT1, a sugarcane gene encoding a MAT1 homolog. Northern blots and in situ hybridization results showed that ScMAT1 was expressed in sugarcane mature leaf, leaf roll and inflorescence, and it was not differentially expressed in any of the other tissues analyzed such us bud and roots. In addition, ScMAT1 was not differentially expressed during different stress conditions and treatment with hormones. In situ hybridization analyses also showed that ScMAT1 was expressed in different cell types during leaf development. In order to identify proteins that interact with ScMAT1, a yeast two hybrid assay with ScMAT1 as bait was used to screen a sugarcane leaf cDNA library. The screening of yeast two hybrids yielded 14 positive clones. One of them is a cytochrome p450 family protein involved in oxidative degradation of toxic compounds. Other clones isolated are also related to plant responses to stress. To determine the subcellular localization of ScMAT1, a ScMAT1-GFP fusion was assayed in onion epidermal cell and the fluorescence was localized to the nucleus, in agreement with the putative role of ScMAT1 as a basal transcription factor.284663672Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [01/07546-1

Bacillus subtilis M4 decreases plant susceptibility towards fungal pathogens by increasing host resistance associated with differential gene expression

Results presented in this paper describe the ability of Bacillus subtilis strain M4 to reduce disease incidence caused by Colletotrichum lagenarium and Pythium aphanidermatum on cucumber and tomato, respectively. Disease protection in both pathosystems was most probably due to induction of resistance in the host plant since experiments were designed in order to avoid any direct contact between the biocontrol agent and the pathogen. Pre-inoculation with strain M4 thus sensitised both plants to react more efficiently to subsequent pathogen infection. In cucumber, the use of endospores provided a disease control level similar to that obtained with vegetative cells. In contrast, a mixture of lipopeptides from the surfactin, iturin and fengycin families showed no resistance-inducing potential. Interestingly, treatment with strain M4 was also associated with significant changes in gene transcription in the host plant as revealed by cDNA-AFLP analyses. Several AFLP fragments corresponded to genes not expressed in control plants and specifically induced by the Bacillus treatment. In support to the macroscopic protective effect, this differential accumulation of mRNA also illustrates the plant reaction following perception of strain M4, and constitutes one of the very first examples of defence-associated modifications at the transcriptional level elicited by a non-pathogenic bacterium in a host plant