Arabidopsis thaliana plastidial methionine sulfoxide reductases B,MSRBs, account for most leaf peptide MSR activity and are essential for growth under environmental constraints through a role in the preservation of photosystem antennae

Methionine oxidation to methionine sulfoxide (MetSO) is reversed by two types of methionine sulfoxide reductases (MSRs), A and B, specific to MetSO S‐ and R‐diastereomers, respectively. Two MSRB isoforms, MSRB1 and MSRB2, are present in chloroplasts of Arabidopsis thaliana. To assess their physiological role, we characterized Arabidopsis mutants knockout for the expression of MSRB1, MSRB2 or both genes. Measurements of MSR activity in leaf extracts revealed that the two plastidial MSRB enzymes account for the major part of leaf peptide MSR capacity. Under standard conditions of light and temperature, plants lacking one or both plastidial MSRBs do not exhibit any phenotype, regarding growth and development. In contrast, we observed that the concomitant absence of both proteins results in a reduced growth for plants cultivated under high light or low temperature. In contrast, double mutant lines restored for MSRB2 expression display no phenotype. Under environmental constraints, the MetSO level in leaf proteins is higher in plants lacking both plastidial MSRBs than in Wt plants. The absence of plastidial MSRBs is associated with an increased chlorophyll a/b ratio, a reduced content of Lhca1 and Lhcb1 proteins and an impaired photosynthetic performance. Finally, we show that MSRBs are able to use as substrates, oxidized cpSRP43 and cpSRP54, the two main components involved in the targeting of Lhc proteins to the thylakoids. We propose that plastidial MSRBs fulfil an essential function in maintaining vegetative growth of plants during environmental constraints, through a role in the preservation of photosynthetic antennae.     

Anti-angiogenesis and anti-tumor activity of recombinant anginex

Anginex, a synthetic 33-mer angiostatic peptide, specifically inhibits vascular endothelial cell proliferation and migration along with induction of apoptosis in endothelial cells. Here we report on the in vivo characterization of recombinant anginex and use of the artificial anginex gene for gene therapy approaches. Tumor growth of human MA148 ovarian carcinoma in athymic mice was inhibited by 80% when treated with recombinant anginex. Histological analysis of the tumors showed an approximate 2.5-fold reduction of microvessel density, suggesting that angiogenesis inhibition is the cause of the anti-tumor effect. Furthermore, there was a significant correlation between the gene expression patterns of 16 angiogenesis-related factors after treatment with both recombinant and synthetic anginex. To validate the applicability of the anginex gene for gene therapy, stable transfectants of murine B16F10 melanoma cells expressing recombinant anginex were made. Supernatants of these cells inhibited endothelial cell proliferation in vitro. Furthermore, after subcutaneous injection of these cells in C57BL/6 mice, an extensive delay in tumor growth was observed. These data show that the artificial anginex gene can be used to produce a recombinant protein with similar activity as its synthetic counterpart and that the gene can be applied in gene therapy approaches for cancer treatment.     

Glutathione peroxidase-1 deficiency augments proinflammatory cytokine-induced redox signaling and human endothelial cell activation

Glutathione peroxidase-1 (GPx-1) is a crucial antioxidant enzyme, the deficiency of which promotes atherogenesis. Accordingly, we examined the mechanisms by which GPx-1 deficiency enhances endothelial cell activation and inflammation. In human microvascular endothelial cells, we found that GPx-1 deficiency augments intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression by redox-dependent mechanisms that involve NFκB. Suppression of GPx-1 enhanced TNF-α-induced ROS production and ICAM-1 expression, whereas overexpression of GPx-1 attenuated these TNF-α-mediated responses. GPx-1 deficiency prolonged TNF-α-induced IκBα degradation and activation of ERK1/2 and JNK. JNK or NFκB inhibition attenuated TNF-α induction of ICAM-1 and VCAM-1 expression in GPx-1-deficient and control cells, whereas ERK1/2 inhibition attenuated only VCAM-1 expression. To analyze further signaling pathways involved in GPx-1-mediated protection from TNF-α-induced ROS, we performed microarray analysis of human microvascular endothelial cells treated with TNF-α in the presence and absence of GPx-1. Among the genes whose expression changed significantly, dual specificity phosphatase 4 (DUSP4), encoding an antagonist of MAPK signaling, was down-regulated by GPx-1 suppression. Targeted DUSP4 knockdown enhanced TNF-α-mediated ERK1/2 pathway activation and resulted in increased adhesion molecule expression, indicating that GPx-1 deficiency may augment TNF-α-mediated events, in part, by regulating DUSP4.     

Invasive grass density negatively impacts chaparral seedling establishment

Type conversion from native shrubland to invasive annual grassland is on the rise due to global change factors such as prolonged drought and increasing fire frequency. Efforts to restore chaparral ecosystems are limited by current understanding of competitive interactions between shrub seedlings and invasive grasses as well as soil moisture requirements of chaparral seedlings. We set up a restoration experiment where we out-planted Adenostoma fasciculatum seedlings, manipulated invasive grass density, monitored soil moisture at two depths, and tracked seedling survival and biomass. We found that higher invasive grass cover was associated with higher rates of seedling mortality but found no difference in biomass per surviving plant. Soil moisture was higher at 15 cm under the 100% weeded treatment than the 50% weeded and control treatments during January. Lower invasive cover resulted in higher richness of annual native plant species, as plots with 100% invasive removal had higher richness than 50% removal and unplanted control plots. Future restoration efforts in the chaparral will likely be more successful in increasing initial seedling establishment if invasive grass removal is included.     

Feedbacks of Soil Inoculum of Mycorrhizal Fungi Altered by N Deposition on the Growth of a Native Shrub and an Invasive Annual Grass

Anthropogenic nitrogen (N) deposition causes shifts in vegetation types as well as species composition of arbuscular mycorrhizal (AM) fungi and other soil microorganisms. A greenhouse experiment was done to determine whether there are feedbacks between N-altered soil inoculum and growth of a dominant native shrub and an invasive grass species in southern California. The region is experiencing large-scale loss of Artemisia californica shrublands and replacement by invasive annual grasses under N deposition. Artemisia californica and Bromus madritensis ssp. rubens were grown with soil inoculum from experimental plots in a low N deposition site that had (1) N-fertilized and (2) unfertilized soil used for inoculum, as well as (3) high-N soil inoculum from a site exposed to atmospheric N deposition for four decades. All treatments plus a nonmycorrhizal control were given two levels of N fertilizer solution. A. californica biomass was reduced by each of the three inocula compared to uninoculated controls under at least one of the two N fertilizer solutions. The␣inoculum from the N-deposition site caused the greatest growth depressions. By contrast, B.␣madritensis biomass increased with each of the three inocula under at least one, or both, of the N solutions. The different growth responses of the two plant species may be related to the types of AM fungal colonization. B. madritensis was mainly colonized by a fine mycorrhizal endophyte, while A. californica had primarily coarse endophytes. Furthermore, A. californica had a high level of septate, nonmycorrhizal root endophytes, while B. madritensis overall had low levels of these endophytes. The negative biomass response of A. californica seedlings to high N-deposition inoculum may in part explain its decline; a microbially-mediated negative feedback may occur in this system that causes poor␣seedling growth and establishment of A.␣californica in sites subject to N deposition and B. madritensis invasion.     

Blockade but Not Overexpression of the Junctional Adhesion Molecule C Influences Virus-Induced Type 1 Diabetes in Mice

Type 1 diabetes (T1D) results from the autoimmune destruction of insulin-producing beta-cells in the pancreas. Recruitment of inflammatory cells is prerequisite to beta-cell-injury. The junctional adhesion molecule (JAM) family proteins JAM-B and JAM–C are involved in polarized leukocyte transendothelial migration and are expressed by vascular endothelial cells of peripheral tissue and high endothelial venules in lympoid organs. Blocking of JAM-C efficiently attenuated cerulean-induced pancreatitis, rheumatoid arthritis or inflammation induced by ischemia and reperfusion in mice. In order to investigate the influence of JAM-C on trafficking and transmigration of antigen-specific, autoaggressive T-cells, we used transgenic mice that express a protein of the lymphocytic choriomeningitis virus (LCMV) as a target autoantigen in the β-cells of the islets of Langerhans under the rat insulin promoter (RIP). Such RIP-LCMV mice turn diabetic after infection with LCMV. We found that upon LCMV-infection JAM-C protein was upregulated around the islets in RIP-LCMV mice. JAM-C expression correlated with islet infiltration and functional beta-cell impairment. Blockade with a neutralizing anti-JAM-C antibody reduced the T1D incidence. However, JAM-C overexpression on endothelial cells did not accelerate diabetes in the RIP-LCMV model. In summary, our data suggest that JAM-C might be involved in the final steps of trafficking and transmigration of antigen-specific autoaggressive T-cells to the islets of Langerhans.