Dual localization of plant glutamate receptor AtGLR3.4 to plastids and plasmamembrane

Bioinformatic approaches have allowed the identification in Arabidopsis thaliana of twenty genes encoding for homologues of animal ionotropic glutamate receptors (iGLRs). Some of these putative receptor proteins, grouped into three subfamilies, have been located to the plasmamembrane, but their possible location in organelles has not been investigated so far. In the present work we provide multiple evidence for the plastid localization of a glutamate receptor, AtGLR3.4, in Arabidopsis and tobacco. Biochemical analysis was performed using an antibody shown to specifically recognize both the native protein in Arabidopsis and the recombinant AtGLR3.4 fused to YFP expressed in tobacco. Western blots indicate the presence of AtGLR3.4 in both the plasmamembrane and in chloroplasts. In agreement, in transformed Arabidopsis cultured cells as well as in agroinfiltrated tobacco leaves, AtGLR3.4::YFP is detected both at the plasmamembrane and at the plastid level by confocal microscopy. The photosynthetic phenotype of mutant plants lacking AtGLR3.4 was also investigated. These results identify for the first time a dual localization of a glutamate receptor, revealing its presence in plastids and chloroplasts and opening the way to functional studies.     

Regulation of electron transport in microalgae

Unicellular algae are characterized by an extreme flexibility with respect to their responses to environmental constraints. This flexibility probably explains why microalgae show a very high biomass yield, constitute one of the major contributors to primary productivity in the oceans and are considered a promising choice for biotechnological applications. Flexibility results from a combination of several factors including fast changes in the light-harvesting apparatus and a strong interaction between different metabolic processes (e.g. respiration and photosynthesis), which all take place within the same cell. Microalgae are also capable of modifying their photosynthetic electron flow capacity, by changing its maximum rate and/or by diverting photogenerated electrons towards different sinks depending on their growth status. In this review, we will focus on the occurrence and regulation of alternative electron flows in unicellular algae and compare data obtained in these systems with those available in vascular plants. This article is part of a Special Issue entitled: Regulation of Electron Transport in Chloroplasts.     

Incidence,survival and mortality rates of stage-specific bladder cancer in United States: A trend analysis

PurposeTo examine the overall and stage-specific age-adjusted incidence, 5-year survival and mortality rates of bladder cancer (BCa) in the United States, between 1973 and 2009.Materials and methodsA total of 148,315 BCa patients were identified in the Surveillance, Epidemiology and End Results database, between years 1973 and 2009. Incidence, mortality, and 5-year cancer-specific survival rates were calculated. Temporal trends were quantified using the estimated annual percentage change (EAPC) and linear regression models. All analyses were stratified according to disease stage, and further examined according to sex, race, and age groups.ResultsIncidence rate of BCa increased from 21.0 to 25.5/100,000 person-years between 1973 and 2009. Stage-specific analyses revealed an increase incidence for localized stage: 15.4–20.2 (EAPC: +0.5%, p < 0.001) and distant stage: 0.5–0.8 (EAPC: +0.7%, p = 0.001). Stage-specific 5-year survival rates increased for all stages, except for distant disease. No significant changes in mortality were recorded among localized (EAPC: ?0.2%, p = 0.1) and regional stage (EAPC: ?0.1%, p = 0.5). An increase in mortality rates was observed among distant stage (EAPC: +1.0%, p = 0.005). Significant variations in incidence and mortality were recorded when estimates were stratified according to sex, race, and age groups.DiscussionAlbeit statistically significant, virtually all changes in incidence and mortality were minor, and hardly of any clinical importance. Little or no change in BCa cancer control outcomes has been achieved during the study period.     

The PPE Domain of PPE17 Is Responsible for Its Surface Localization and Can Be Used to Express Heterologous Proteins on the Mycobacterial Surface

PPE represent a peculiar family of mycobacterial proteins characterized by a 180 aminoacids conserved N-terminal domain. Several PPE genes are co-transcribed with a gene encoding for a protein belonging to another family of mycobacterial specific proteins named PE. Only one PE-PPE couple has been extensively characterized so far (PE25-PPE41) and it was shown that these two proteins form a heterodimer and that this interaction is essential for PPE41 stability and translocation through the mycobacterial cell wall. In this study we characterize the PE11-PPE17 couple. In contrast with what was found for PE25-PPE41, we show that PPE17 is not secreted but surface exposed. Moreover, we demonstrate that the presence of PE11 is not necessary for PPE17 stability or for its localization on the mycobacterial surface. Finally, we show that the PPE domain of PPE17 targets the mycobacterial cell wall and that this domain can be used as a fusion partner to expose heterologous proteins on the mycobacterial surface.     

Association of Soluble Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) with Central Adiposity and Low-Density Lipoprotein Cholesterol

Objective

Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL), in addition to having a prognostic value in patients with cardiovascular disease, seems to interact with adiposity, insulin resistance and other cardiovascular risk factors. However, the results of previous clinical studies, focused on the association of TRAIL with selected metabolic or anthropometric indices were inconclusive. The aim of this study was to further investigate how soluble TRAIL concentrations independently correlate with major cardiovascular risk factors, including lipid, glycemic and anthropometric features.

Materials/Methods

We examined the associations between serum soluble TRAIL concentrations, measured by ELISA, and lipid, glycemic and anthropometric features in 199 subjects recruited at our Metabolic Outpatient Clinic.

Results

Soluble TRAIL concentrations had a significant and direct correlation with total cholesterol (p = 0.046), LDL-cholesterol (p = 0.032), triglycerides (p = 0.01), body mass index (p = 0.046), waist circumference (p = 0.008), fat mass (p = 0.056) and insulin (p = 0.046) and an inverse correlation with HDL-cholesterol (p = 0.02). In multivariable regression analyses adjusted for potential confounders (age, gender, C-reactive protein, HDL-cholesterol, triglycerides, waist circumference, and insulin), TRAIL levels continued to have an independent correlation with LDL-cholesterol and waist circumference (r² = 0.04).

Conclusions

Serum TRAIL levels were weakly but significantly and independently associated with waist circumference, a marker of visceral adiposity, and with LDL-cholesterol. Further studies are needed to clarify the biological basis of these relationships.     

New Insights on Cytological and Metabolic Features of Ostreopsis cf. ovata Fukuyo (Dinophyceae): A Multidisciplinary Approach

The harmful dinoflagellate Ostreopsis cf. ovata has been causing toxic events along the Mediterranean coasts and other temperate and tropical areas, with increasing frequency during the last decade. Despite many studies, important biological features of this species are still poorly known. An integrated study, using different microscopy and molecular techniques, Raman microspectroscopy and high resolution liquid chromatography-mass spectrometry (HR LC-MS), was undertaken to elucidate cytological aspects, and identify main metabolites including toxins. The species was genetically identified as O. cf. ovata, Atlantic-Mediterranean clade. The ultrastructural results show unique features of the mucilage network abundantly produced by this species to colonize benthic substrates, with a new role of trichocysts, never described before. The amorphous polysaccharidic component of mucilage appears to derive from pusule fibrous material and mucocysts. In all stages of growth, the cells show an abundant production of lipids. Different developmental stages of chloroplasts are found in the peripheral cytoplasm and in the centre of cell. In vivo Raman microspectroscopy confirms the presence of the carotenoid peridinin in O. cf. ovata, and detects in several specimen the abundant presence of unsaturated lipids structurally related to docosahexaenoic acid. The HR LC-MS analysis reveals that ovatoxin-a is the predominant toxin, together with decreasing amounts of ovatoxin-b, -d/e, -c and putative palytoxin. Toxins concentration on a per cell basis increases from exponential to senescent phase. The results suggest that benthic blooms of this species are probably related to features such as the ability to create a unique mucilaginous sheath covering the sea bottom, associated with the production of potent toxins as palytoxin-like compounds. In this way, O. cf. ovata may be able to rapidly colonize benthic substrates outcompeting other species.     

A Fasting Insulin–Raising Allele at IGF1 Locus Is Associated with Circulating Levels of IGF-1 and Insulin Sensitivity

Background

A meta-analysis of genome-wide data reported the discovery of the rs35767 polymorphism near IGF1 with genome-wide significant association with fasting insulin levels. However, it is unclear whether the effects of this polymorphism on fasting insulin are mediated by a reduced insulin sensitivity or impaired insulin clearance. We investigated the effects of the rs35767 polymorphism on circulating IGF-1 levels, insulin sensitivity, and insulin clearance.

Methodology/Principal Findings

Two samples of adult nondiabetic white Europeans were studied. In sample 1 (n=569), IGF-1 levels were lower in GG genotype carriers compared with A allele carriers (190±77 vs. 218±97 ng/ml, respectively; P=0.007 after adjusting for age, gender, and BMI). Insulin sensitivity assessed by euglycaemic-hyperinsulinemic clamp was lower in GG genotype carriers compared with A allele carriers (8.9±4.1 vs. 10.1±5.1 mg x Kg^-1 free fat mass x min^-1, respectively; P=0.03 after adjusting for age, gender, and BMI). The rs35767 polymorphism did not show significant association with insulin clearance. In sample 2 (n=859), IGF-1 levels were lower in GG genotype carriers compared with A allele carriers (155±60 vs. 164±63 ng/ml, respectively; P=0.02 after adjusting for age, gender, and BMI). Insulin sensitivity, as estimated by the HOMA index, was lower in GG genotype carriers compared with A allele carriers (2.8±2.2 vs. 2.5±1.3, respectively; P=0.03 after adjusting for age, gender, and BMI).

Conclusion/Significance

The rs35767 polymorphism near IGF1 was associated with circulating IGF-1 levels, and insulin sensitivity with carriers of the GG genotype exhibiting lower IGF-1 concentrations and insulin sensitivity as compared with subjects carrying the A allele.     

Does the environment around the H-cluster allow coordination of the pendant amine to the catalytic iron center in [FeFe] hydrogenases? Answers from theory

[FeFe] hydrogenases are H₂-evolving enzymes that feature a diiron cluster in their active site (the [2Fe]_H cluster). One of the iron atoms has a vacant coordination site that directly interacts with H₂, thus favoring its splitting in cooperation with the secondary amine group of a neighboring, flexible azadithiolate ligand. The vacant site is also the primary target of the inhibitor O₂. The [2Fe]_H cluster can span various redox states. The active-ready form (H_ox) attains the Fe^IIFe^I state. States more oxidized than H_ox were shown to be inactive and/or resistant to O₂. In this work, we used density functional theory to evaluate whether azadithiolate-to-iron coordination is involved in oxidative inhibition and protection against O₂, a hypothesis supported by recent results on biomimetic compounds. Our study shows that Fe–N(azadithiolate) bond formation is favored for an Fe^IIFe^II active-site model which disregards explicit treatment of the surrounding protein matrix, in line with the case of the corresponding Fe^IIFe^II synthetic system. However, the study of density functional theory models with explicit inclusion of the amino acid environment around the [2Fe]_H cluster indicates that the protein matrix prevents the formation of such a bond. Our results suggest that mechanisms other than the binding of the azadithiolate nitrogen protect the active site from oxygen in the so-called H _ox ^inact state.