Mevalonosomes: specific vacuoles containing the mevalonate pathway in Plocamium brasiliense cortical cells (Rhodophyta)

This paper has identified, for the first time in a member of the Rhodophyta, a vacuolar organelle containing enzymes that are involved in the mevalonate pathway—an important step in red algal isoprenoid biosynthesis. These organelles were named mevalonosomes (Mev) and were found in the cortical cells (CC) of Plocamium brasiliense, a marine macroalgae that synthesizes several halogenated monoterpenes. P. brasiliense specimens were submitted to a cytochemical analysis of the activity of the 3‐hydroxy‐3‐methylglutaryl‐CoA synthase (HMGS). Using transmission electron microscopy (TEM), we confirmed the presence of HMGS activity within the Mev. Because HMGS is necessary for the biosynthesis of halogenated monoterpenes, we isolated a hexanic fraction (HF) rich in halogenated monoterpenes from P. brasiliense that contained a pentachlorinated monoterpene as a major metabolite. Because terpenes are often related to chemical defense, the antifouling (AF) activity of pentachlorinated monoterpene was tested. We found that the settlement of the mussel Perna perna was reduced by HF treatment (2.25 times less than control; 40% and 90% of fouled surface, respectively; P = 0.001; F_9,9 = 1.13). The HF (at 10 μg · mL^?1) also inhibited three species of fouling microalgae (Chlorarachnion reptans, Cylindrotheca cloisterium, and Exanthemachrysis gayraliae), while at a higher concentration (50 μg · mL^?1), it inhibited the bacteria Halomonas marina, Polaribacter irgensii, Pseudoalteromonas elyakovii, Shewanella putrefaciens, and Vibrio aestuarianus. The AF activity of P. brasiliense halogenated monoterpenes and the localization of HMGS activity inside Mev suggest that this cellular structure found in CC may play a role in thallus protection against biofouling.     

Physical association of the NB-LRR resistance protein Rx with a Ran GTPase-activating protein is required for extreme resistance to Potato virus X

Nucleotide binding leucine-rich repeat (NB-LRR) proteins play an important role in plant and mammalian innate immunity. In plants, these resistance proteins recognize specific pathogen-derived effector proteins. Recognition subsequently triggers a rapid and efficient defense response often associated with the hypersensitive response and other poorly understood processes that suppress the pathogen. To investigate mechanisms associated with the activation of disease resistance responses, we investigated proteins binding to the potato (Solanum tuberosum) NB-LRR protein Rx that confers extreme resistance to Potato virus X (PVX) in potato and Nicotiana benthamiana. By affinity purification experiments, we identified an endogenous N. benthamiana Ran GTPase-Activating Protein2 (RanGAP2) as an Rx-associated protein in vivo. Further characterization confirmed the specificity of this interaction and showed that the association occurs through their N-terminal domains. By specific virus-induced gene silencing of RanGAP2 in N. benthamiana carrying Rx, we demonstrated that this interaction is required for extreme resistance to PVX and suggest that RanGAP2 is part of the Rx signaling complex. These results implicate RanGAP-mediated cellular mechanisms, including nucleocytoplasmic trafficking, in the activation of disease resistance.     

Phylogeography, population dynamics, and molecular evolution of European bat lyssaviruses

European bat lyssaviruses types 1 and 2 (EBLV-1 and EBLV-2) are widespread in Europe, although little is known of their evolutionary history. We undertook a comprehensive sequence analysis to infer the selection pressures, rates of nucleotide substitution, age of genetic diversity, geographical origin, and population growth rates of EBLV-1. Our study encompassed data from 12 countries collected over a time span of 35 years and focused on the glycoprotein (G) and nucleoprotein (N) genes. We show that although the two subtypes of EBLV-1--EBLV-1a and EBLV-1b--have both grown at a low exponential rate since their introduction into Europe, they have differing population structures and dispersal patterns. Furthermore, there were strong constraints against amino acid change in both EBLV-1 and EBLV-2, as reflected in a low ratio of nonsynonymous to synonymous substitutions per site, particularly in EBLV-1b. Our inferred rate of nucleotide substitution in EBLV-1, approximately 5 x 10(-5) substitutions per site per year, was also one of the lowest recorded for RNA viruses and implied that the current genetic diversity in the virus arose 500 to 750 years ago. We propose that the slow evolution of EBLVs reflects their distinctive epidemiology in bats, where they occupy a relatively stable fitness peak.     

1,5-Anhydroglucitol predicts CKD progression in macroalbuminuric diabetic kidney disease: results from non-targeted metabolomics

Introduction

Metabolomics allows exploration of novel biomarkers and provides insights on metabolic pathways associated with disease. To date, metabolomics studies on CKD have been largely limited to Caucasian populations and have mostly examined surrogate end points.

Objective

In this study, we evaluated the role of metabolites in predicting a primary outcome defined as dialysis need, doubling of serum creatinine or death in Brazilian macroalbuminuric DKD patients.

Methods

Non-targeted metabolomics was performed on plasma from 56 DKD patients. Technical triplicates were done. Metabolites were identified using Agilent Fiehn GC/MS Metabolomics and NIST libraries (Agilent MassHunter Work-station Quantitative Analysis, version B.06.00). After data cleaning, 186 metabolites were left for analyses.

Results

During a median follow-up time of 2.5 years, the PO occurred in 17 patients (30.3%). In non-parametric testing, 13 metabolites were associated with the PO. In univariate Cox regression, only 1,5-anhydroglucitol (HR 0.10; 95% CI 0.01–0.63, p?=?.01), norvaline and l-aspartic acid were associated with the PO. After adjustment for baseline renal function, 1,5-anhydroglucitol (HR 0.10; 95% CI 0.02–0.63, p?=?.01), norvaline (HR 0.01; 95% CI 0.001–0.4, p?=?.01) and aspartic acid (HR 0.12; 95% CI 0.02–0.64, p?=?.01) remained significantly and inversely associated with the PO.

Conclusion

Our results show that lower levels of 1,5-anhydroglucitol, norvaline and l-aspartic acid are associated with progression of macroalbuminuric DKD. While norvaline and l-aspartic acid point to interesting metabolic pathways, 1,5-anhydroglucitol is of particular interest since it has been previously shown to be associated with incident CKD. This inverse biomarker of hyperglycemia should be further explored as a new tool in DKD.

     

An unusual presentation of paracoccidioidomycosis in an AIDS patient: A case report

A case of paracoccidioidomycosis presenting as a solitary pulmonary nodular lesion in a patient with acquired immunodeficiency syndrome (AIDS) is presented. This case illustrates that restricted lung lesions can also be found and diagnosed in immunodeficient patients. This revised version was published online in June 2006 with corrections to the Cover Date.     

Metalloproteomics Approach to Analyze Mercury in Breast Milk and Hair Samples of Lactating Women in Communities of the Amazon Basin,Brazil

Mercury is a potentially toxic element that is present in the environment of the Brazilian Amazon and is responsible for adverse health effects in humans. This study sought to assess possible protein biomarkers of mercury exposure in breast milk samples from lactating women in the Madeira and Negro Rivers in the Brazilian Amazon. The mercury content of hair samples of lactating women was determined, and the proteome of breast milk samples was obtained using two-dimensional electrophoresis after protein precipitation with acetone. Mercury measurements of protein spots obtained via protein fractionation were performed by graphite furnace atomic absorption spectrometry (GFAAS), and it was observed that mercury is linked to proteins with molecular masses in the range of 14–26 kDa. The total mercury concentration was also determined by GFAAS in unprocessed milk, lyophilized milk, and protein pellets, with the purpose of determining the mercury mass balance in relation to the concentration of this element in milk and pellets. Approximately 85 to 97% of mercury present in the lyophilized milk from samples of lactating women of the Madeira River is bound in the protein fraction. From lactating women of the Negro River, approximately 49% of the total mercury is bound in the protein fraction, and a difference of 51% is bound in the lipid fraction.     

Inflammatory profile in X‐linked adrenoleukodystrophy patients: Understanding disease progression

X‐linked adrenoleukodystrophy (X‐ALD) is an inherited disease characterized by progressive inflammatory demyelization in the brain, adrenal insufficiency, and an abnormal accumulation of very long chain fatty acids (VLCFA) in tissue and body fluids. Considering that inflammation might be involved in pathophysiology of X‐ALD, we aimed to investigate pro‐ and anti‐inflammatory cytokines in plasma from three different male phenotypes (CCER, AMN, and asymptomatic individuals). Our results showed that asymptomatic patients presented increased levels of pro‐inflammatory cytokines IL‐1β, IL‐2, IL‐8, and TNF‐α and the last one was also higher in AMN phenotype. Besides, asymptomatic patients presented higher levels of anti‐inflammatory cytokines IL‐4 and IL‐10. AMN patients presented higher levels of IL‐2, IL‐5, and IL‐4. We might hypothesize that inflammation in X‐ALD is related to plasmatic VLCFA concentration, since there were positive correlations between C26:0 plasmatic levels and pro‐inflammatory cytokines in asymptomatic and AMN patients and negative correlation between anti‐inflammatory cytokine and C24:0/C22:0 ratio in AMN patients. The present work yields experimental evidence that there is an inflammatory imbalance associated Th1, (IL‐2, IL‐6, and IFN‐γ), Th2 (IL‐4 and IL‐10), and macrophages response (TNF‐α and IL‐1β) in the periphery of asymptomatic and AMN patients, and there is correlation between VLCFA plasmatic levels and inflammatory mediators in X‐ALD. Furthermore, we might also speculate that the increase of plasmatic cytokines in asymptomatic patients could be considered an early biomarker of brain damage and maybe also a predictor of disease progression.     

Oxidative Imbalance,Nitrative Stress,and Inflammation in C6 Glial Cells Exposed to Hexacosanoic Acid: Protective Effect of <Emphasis Type="Italic">N</Emphasis>-acetyl-<Emphasis Type="SmallCaps">l</Emphasis>-cysteine,Trolox, and Rosuvastatin

X-linked adrenoleukodystrophy (X-ALD) is an inherited neurometabolic disorder caused by disfunction of the ABCD1 gene, which encodes a peroxisomal protein responsible for the transport of the very long-chain fatty acids from the cytosol into the peroxisome, to undergo β-oxidation. The mainly accumulated saturated fatty acids are hexacosanoic acid (C26:0) and tetracosanoic acid (C24:0) in tissues and body fluids. This peroxisomal disorder occurs in at least 1 out of 20,000 births. Considering that pathophysiology of this disease is not well characterized yet, and glial cells are widely used in studies of protective mechanisms against neuronal oxidative stress, we investigated oxidative damages and inflammatory effects of vesicles containing lecithin and C26:0, as well as the protection conferred by N-acetyl-l-cysteine (NAC), trolox (TRO), and rosuvastatin (RSV) was assessed. It was verified that glial cells exposed to C26:0 presented oxidative DNA damage (measured by comet assay and endonuclease III repair enzyme), enzymatic oxidative imbalance (high catalase activity), nitrative stress [increased nitric oxide (NO) levels], inflammation [high Interleukin-1beta (IL-1β) levels], and induced lipid peroxidation (increased isoprostane levels) compared to native glial cells without C26:0 exposure. Furthermore, NAC, TRO, and RSV were capable to mitigate some damages caused by the C26:0 in glial cells. The present work yields experimental evidence that inflammation, oxidative, and nitrative stress may be induced by hexacosanoic acid, the main accumulated metabolite in X-ALD, and that antioxidants might be considered as an adjuvant therapy for this severe neurometabolic disease.     

Biochemical and Structural Study of the Atypical Acyltransferase Domain from the Mycobacterial Polyketide Synthase Pks13

Pks13 is a type I polyketide synthase involved in the final biosynthesis step of mycolic acids, virulence factors, and essential components of the Mycobacterium tuberculosis envelope. Here, we report the biochemical and structural characterization of a 52-kDa fragment containing the acyltransferase domain of Pks13. This fragment retains the ability to load atypical extender units, unusually long chain acyl-CoA with a predilection for carboxylated substrates. High resolution crystal structures were determined for the apo, palmitoylated, and carboxypalmitoylated forms. Structural conservation with type I polyketide synthases and related fatty-acid synthases also extends to the interdomain connections. Subtle changes could be identified both in the active site and in the upstream and downstream linkers in line with the organization displayed by this singular polyketide synthase. More importantly, the crystallographic analysis illustrated for the first time how a long saturated chain can fit in the core structure of an acyltransferase domain through a dedicated channel. The structures also revealed the unexpected binding of a 12-mer peptide that might provide insight into domain-domain interaction.     

Food Selection of Cafeteria Diet Affects Memory Dysfunction Related to Obesity

Neurochemical Research - Cafeteria diet (CAF) mimics human Western diet and has been used in animal models to study obesity. The purpose of this study is to demonstrate that our CAF model induces...