Autophagy in Spinocerebellar ataxia type 2, a dysregulated pathway,and a target for therapy

Spinocerebellar ataxia type 2 (SCA2) is an incurable and genetic neurodegenerative disorder. The disease is characterized by progressive degeneration of several brain regions, resulting in severe motor and non-motor clinical manifestations. The mutation causing SCA2 disease is an abnormal expansion of CAG trinucleotide repeats in the ATXN2 gene, leading to a toxic expanded polyglutamine segment in the translated ataxin-2 protein. While the genetic cause is well established, the exact mechanisms behind neuronal death induced by mutant ataxin-2 are not yet completely understood. Thus, the goal of this study is to investigate the role of autophagy in SCA2 pathogenesis and investigate its suitability as a target for therapeutic intervention. For that, we developed and characterized a new striatal lentiviral mouse model that resembled several neuropathological hallmarks observed in SCA2 disease, including formation of aggregates, neuronal marker loss, cell death and neuroinflammation. In this new model, we analyzed autophagic markers, which were also analyzed in a SCA2 cellular model and in human post-mortem brain samples. Our results showed altered levels of SQSTM1 and LC3B in cells and tissues expressing mutant ataxin-2. Moreover, an abnormal accumulation of these markers was detected in SCA2 patients’ striatum and cerebellum. Importantly, the molecular activation of autophagy, using the compound cordycepin, mitigated the phenotypic alterations observed in disease models. Overall, our study suggests an important role for autophagy in the context of SCA2 pathology, proposing that targeting this pathway could be a potential target to treat SCA2 patients.Subject terms: Diseases of the nervous system, Molecular neuroscience     

N‐glycan maturation mutants in Lotus japonicus for basic and applied glycoprotein research

Studies of protein N‐glycosylation are important for answering fundamental questions on the diverse functions of glycoproteins in plant growth and development. Here we generated and characterised a comprehensive collection of Lotus japonicusLORE1 insertion mutants, each lacking the activity of one of the 12 enzymes required for normal N‐glycan maturation in the glycosylation machinery. The inactivation of the individual genes resulted in altered N‐glycan patterns as documented using mass spectrometry and glycan‐recognising antibodies, indicating successful identification of null mutations in the target glyco‐genes. For example, both mass spectrometry and immunoblotting experiments suggest that proteins derived from the α1,3‐fucosyltransferase (Lj3fuct) mutant completely lacked α1,3‐core fucosylation. Mass spectrometry also suggested that the Lotus japonicus convicilin 2 was one of the main glycoproteins undergoing differential expression/N‐glycosylation in the mutants. Demonstrating the functional importance of glycosylation, reduced growth and seed production phenotypes were observed for the mutant plants lacking functional mannosidase I, N‐acetylglucosaminyltransferase I, and α1,3‐fucosyltransferase, even though the relative protein composition and abundance appeared unaffected. The strength of our N‐glycosylation mutant platform is the broad spectrum of resulting glycoprotein profiles and altered physiological phenotypes that can be produced from single, double, triple and quadruple mutants. This platform will serve as a valuable tool for elucidating the functional role of protein N‐glycosylation in plants. Furthermore, this technology can be used to generate stable plant mutant lines for biopharmaceutical production of glycoproteins displaying relative homogeneous and mammalian‐like N‐glycosylation features.     

Pollen analysis of honeys from the northwest of Argentina: Province of Jujuy

The palynological characterisation of 157 honey samples from three northwest regions of Argentina (Prepuna, Yungas and Chaco) are presented to determine their botanical origin and species associations to be able to define their geographic origin. Samples were harvested during 2003–2011 and processed by means of melissopalynological conventional techniques. One-hundred and nine pollen types were identified. Representative pollen types with a frequency of occurrence greater than 50% in descending order of importance are: Salix humboldtiana, Allophylus edulis, Baccharis, Solanaceae, Eucalyptus, Schinus, Brassicaceae, Papilionoideae, Celtis, Scutia/Condalia-type and Parapiptadenia excelsa. The most important monofloral honeys are from the following: Salix humboldtiana, Scutia/Condalia-type, Allophylus edulis, Baccharis, Blepharocalyx salicifolius, Gleditsia amorphoides, Myrtaceae, Sicyos, Ziziphus mistol, Schinopsis-type, Agonandra excelsa, Anadenathera colubrina, Mimosa, all of them native species, and among introduced species are Eucalyptus, Citrus and Tithonia. Three apicultural zones and their corresponding pollen association indicators were determined: Zone I, Prepuna: Arquita trichocarpa, Prosopis ferox, Schinus areira, Baccharis, Buddleja and Mutisieae; Zone II, Yungas: Myrtaceae, Parapiptadenia excelsa, Baccharis, Salix humboldtiana, Allophylus edulis, Scutia/Condalia-type and Zanthoxylum coco; Zone III, transitional area Yungas-Chaco: Prosopis, Salix humboldtiana, Schinus, Anadenanthera colubrina and Allophylus edulis.     

Chronic Stress Causes Sex-Specific and Structure-Specific Alterations in Mitochondrial Respiratory Chain Activity in Rat Brain

Chronic restraint stress (CRS) induces a variety of changes in brain function, some of which are mediated by glucocorticoids. The response to stress occurs in a sex-specific way, and may include mitochondrial and synaptic alterations. The synapse is highly dependent on mitochondrial energy supply, and when mitochondria become dysfunctional, they orchestrate cell death. This study aimed to investigate the CRS effects on mitochondrial respiratory chain activity, as well as mitochondrial potential and mass in cell body and synapses using hippocampus, cortex and striatum of male and female rats. Rats were divided into non-stressed (control) and stressed group (CRS during 40 days). Results showed that CRS increased complex I–III activity in hippocampus. We also observed an interaction between CRS and sex in the striatal complex II activity, since CRS induced a reduction in complex II activity in males, while in females this activity was increased. Also an interaction was observed between stress and sex in cortical complex IV activity, since CRS induced increased activity in females, while it was reduced in males. Glucocorticoid receptor (GR) content in cortex and hippocampus was sexually dimorphic, with female rats presenting higher levels compared to males. No changes were observed in GR content, mitochondrial potential or mass of animals submitted to CRS. It was concluded that CRS induced changes in respiratory chain complex activities, and some of these changes are sex-dependent: these activities are increased in the striatal mitochondria by CRS protocol mainly in females, while in males it is decreased.

     

Tungsten accumulation by highly tolerant marine hydrothermal Sulfitobacter dubius strains carrying a tupBCA cluster

Tungsten (W) has industrial and economic importance, and is in the European Union list of metals with a high supply risk. It is used by living organisms, which transport it into the cell, in the form of tungstate ion (WO₄^2?), using three different ABC-type transporters from the specific W-uptake system coded by tupABC gene cluster. In this study, strains from a collection recovered from deep-sea hydrothermal sediments were selected according to their ability to tolerate metals and to possess the tup genetic determinants. Three multimetal-tolerant strains, Sulfitobacter dubius NA4, As(V)4 and Sb5, were chosen. The strains were able to grow in the presence of high tungsten concentrations and their growth was unaffected by 1 mM tungsten. Moreover, strain Sb5 was able to accumulate up to 52 μg W mg^?1 protein. Their tup genes were shown to be organized as tupBCA, which is not the most usual gene arrangement. All three strains had the classical TupA conserved motif TTTS, comprising a first Thr replaced by a Val, which seems to be a common feature of the genus Sulfitobacter. This study was an important first step in the exploration of new biological strategies for recovering tungsten from natural or anthropogenic W-impacted environments.     

Genomic distribution and estimation of nucleotide diversity in natural populations: perspectives from the collared flycatcher (Ficedula albicollis) genome

Properly estimating genetic diversity in populations of nonmodel species requires a basic understanding of how diversity is distributed across the genome and among individuals. To this end, we analysed whole‐genome resequencing data from 20 collared flycatchers (genome size ≈1.1 Gb; 10.13 million single nucleotide polymorphisms detected). Genomewide nucleotide diversity was almost identical among individuals (mean = 0.00394, range = 0.00384–0.00401), but diversity levels varied extensively across the genome (95% confidence interval for 200‐kb windows = 0.0013–0.0053). Diversity was related to selective constraint such that in comparison with intergenic DNA, diversity at fourfold degenerate sites was reduced to 85%, 3′ UTRs to 82%, 5′ UTRs to 70% and nondegenerate sites to 12%. There was a strong positive correlation between diversity and chromosome size, probably driven by a higher density of targets for selection on smaller chromosomes increasing the diversity‐reducing effect of linked selection. Simulations exploring the ability of sequence data from a small number of genetic markers to capture the observed diversity clearly demonstrated that diversity estimation from finite sampling of such data is bound to be associated with large confidence intervals. Nevertheless, we show that precision in diversity estimation in large outbred population benefits from increasing the number of loci rather than the number of individuals. Simulations mimicking RAD sequencing showed that this approach gives accurate estimates of genomewide diversity. Based on the patterns of observed diversity and the performed simulations, we provide broad recommendations for how genetic diversity should be estimated in natural populations.     

Diagnosis of Helicobacter pylori

The different invasive and noninvasive diagnostic tests for Helicobacter pylori have been applied mainly in emerging countries. Molecular methods have been developed, especially a test for detection of H. pylori and its clarithromycin resistance directly from stools. The long-term effects of eradication on histologic lesions have been studied in a meta-analysis and the prognostic value of post-treatment in gastric mucosa-associated lymphoid tissue lymphoma has been assessed. An operating link for gastritis assessment (the OLGA staging) has also been published. Attempts to simplify the urea breath test protocol have been made, and new stool antigen tests have been proposed and compared to those previously available.     

Evaluation of a novel monoclonal-based antigen-in-stool enzyme immunoassay (Premier Platinum HpSA PLUS) for diagnosis of Helicobacter pylori infection in Vietnamese children

Background: Helicobacter pylori infection is difficult to diagnose in children, especially in developing countries where noninvasive methods such as urea breath test are often not available. We evaluate the sensitivity and specificity of a new monoclonal antibody-based antigen-in-stool enzyme immunoassay (Premier Platinum HpSA PLUS) for diagnosis of H. pylori infection in Vietnamese children.
Materials and Methods: Sensitivity of the antigen-in-stool test was evaluated in 232 children, 3–15 years of age, who were positive for H. pylori infection by culture from biopsies. For evaluation of the specificity 98 children of similar age with nongastrointestinal conditions and who were negative for H. pylori infection by serologic assays were included with blood and stool samples.
Results: Of the 232 culture-positive children, 224 were also positive by Premier Platinum HpSA PLUS. Of the 98 control children, 93 were H. pylori negative also in the stool test. The sensitivity of Premier Platinum HpSA PLUS was thus 96.6% (95% CI 93.3–98.5) and the specificity was 94.9% (95% CI 88.5–98.3).
Conclusions: The findings have demonstrated Premium Platinum HpSA PLUS to be a reliable method for detection of H. pylori infection also in children in our area.     

A new species of Laranda Walker 1869 (Orthoptera: Grylloidea, Phalangopsidae) from remnant patches of the Brazilian Atlantic Forest

The genus Laranda has six described species and is confined to South and Southeast of Brazil. We describe a new species and discuss the biology and distribution of the genus. The new species can be distinguished from its known congeners by the following characteristics: absence of yellow spots on pronotum and base of posterior tibiae; female copulatory papilla: sclerotization in dorsal view forming opposing acute angles, apical lobes narrow and small; male genitalia: pseudepiphallic median process short and wide; pseudepiphallic paramere with apex incurved and ectophallic fold surpassing apex of the parameres. The genus is distributed within the Atlantic Forest biome; the new species is found on tree trunks, as well as on forest leaf litter.     

Structural insights into hydroxycoumarin-induced apoptosis in U-937 cells

In the present study, we sought to establish the effect of diverse structural-related hydroxycoumarins on the proliferation, cytotoxicity, and induction of apoptosis in promonocytic leukemic cells (U-937). The dihydroxylated coumarins, 7,8-dihydroxy-coumarin and esculetin, induced DNA fragmentation as well as characteristic morphological changes of programmed cell death in U-937 cells. With the aim to perform a structure-activity relationship study, the correlation between the physicochemical properties of the molecules and their pro-apoptotic activity was carried out. Results showed that the presence of two adjacent phenolic hydroxyl groups was the most important factor in terms of the SAR. The exposure of leukemic cells to 7,8-dihydroxy-coumarin evoked a phenoxyl radical generation that was detected by electron spin resonance spectroscopy. The present study suggests that reactive oxygen species generation plays a critical role in dihydroxycoumarin-induced apoptosis in U-937 cells. These findings further suggest that these compounds may have a potential therapeutic role in the treatment of hematological malignancies.