Targeting Dyrk1A with AAVshRNA attenuates motor alterations in TgDyrk1A, a mouse model of Down syndrome

Genetic-dissection studies carried out with Down syndrome (DS) murine models point to the critical contribution of Dyrk1A overexpression to the motor abnormalities and cognitive deficits displayed in DS individuals. In the present study we have used a murine model overexpressing Dyrk1A (TgDyrk1A mice) to evaluate whether functional CNS defects could be corrected with an inhibitory RNA against Dyrk1A, delivered by bilateral intrastriatal injections of adeno-associated virus type 2 (AAVshDyrk1A). We report that AAVshDyrk1A efficiently transduced HEK293 cells and primary neuronal cultures, triggering the specific inhibition of Dyrk1A expression. Injecting the vector into the striata of TgDyrk1A mice resulted in a restricted, long-term transduction of the striatum. This gene therapy was found to be devoid of toxicity and succeeded in normalizing Dyrk1A protein levels in TgDyrk1A mice. Importantly, the behavioral studies of the adult TgDyrk1A mice treated showed a reversal of corticostriatal-dependent phenotypes, as revealed by the attenuation of their hyperactive behavior, the restoration of motor-coordination defects, and an improvement in sensorimotor gating. Taken together, the data demonstrate that normalizing Dyrk1A gene expression in the striatum of adult TgDyrk1A mice, by means of AAVshRNA, clearly reverses motor impairment. Furthermore, these results identify Dyrk1A as a potential target for therapy in DS.     

Low metabolic rates in primitive hunters and weaver spiders

The rates of oxygen consumption and carbon dioxide release of primitive hunters and weaver spiders, the Chilean Recluse spider Loxosceles laeta Nicolet (Araneae: Sicariidae) and the Chilean Tiger spider Scytodes globula Nicolet (Araneae: Scytodidae), are analyzed, and their relationship with body mass is studied. The results are compared with the metabolic data available for other spiders. A low metabolic rate is found both for these two species and other primitive hunters and weavers, such as spiders of the families Dysderidae and Plectreuridae. The metabolic rate of this group is lower than in nonprimitive spiders, such as the orb weavers (Araneae: Araneidae). The results reject the proposition of a general relationship for metabolic rate for all land arthropods (related to body mass) and agree with the hypothesis that metabolic rates are affected not only by sex, reproductive and developmental status, but also by ecology and life style, recognizing here, at least in the araneomorph spiders, a group having low metabolism, comprising the primitive hunters and weaver spiders, and another group comprising the higher metabolic rate web building spiders (e.g. orb weavers).     

Reproducibility and Consistency of Proteomic Experiments on Natural Populations of a Non-Model Aquatic Insect

Population proteomics has a great potential to address evolutionary and ecological questions, but its use in wild populations of non-model organisms is hampered by uncontrolled sources of variation. Here we compare the response to temperature extremes of two geographically distant populations of a diving beetle species (Agabus ramblae) using 2-D DIGE. After one week of acclimation in the laboratory under standard conditions, a third of the specimens of each population were placed at either 4 or 27°C for 12 h, with another third left as a control. We then compared the protein expression level of three replicated samples of 2–3 specimens for each treatment. Within each population, variation between replicated samples of the same treatment was always lower than variation between treatments, except for some control samples that retained a wider range of expression levels. The two populations had a similar response, without significant differences in the number of protein spots over- or under-expressed in the pairwise comparisons between treatments. We identified exemplary proteins among those differently expressed between treatments, which proved to be proteins known to be related to thermal response or stress. Overall, our results indicate that specimens collected in the wild are suitable for proteomic analyses, as the additional sources of variation were not enough to mask the consistency and reproducibility of the response to the temperature treatments.     

Glutathione depletion switches nitric oxide neurotrophic effects to cell death in midbrain cultures: implications for Parkinson's disease

Nitric oxide (NO) exerts neurotrophic and neurotoxic effects on dopamine (DA) function in primary midbrain cultures. We investigate herein the role of glutathione (GSH) homeostasis in the neurotrophic effects of NO. Fetal midbrain cultures were pretreated with GSH synthesis inhibitor, l ‐buthionine‐(S,R)‐sulfoximine (BSO), 24 h before the addition of NO donors (diethylamine/nitric oxide‐complexed sodium and S‐nitroso‐N‐acetylpenicillamine) at doses tested previously as neurotrophic. Under these conditions, the neurotrophic effects of NO disappeared and turned on highly toxic. Reduction of GSH levels to 50% of baseline induced cell death in response to neurotrophic doses of NO. Soluble guanylate cyclase (sGC) and cyclic GMP‐dependent protein kinase (PKG) inhibitors protected from cell death for up to 10 h after NO addition; the antioxidant ascorbic acid also protected from cell death but its efficacy decreased when it was added after NO treatment (40% protection 2 h after NO addition). The pattern of cell death was characterized by an increase in chromatin condensed cells with no DNA fragmentation and with breakdown of plasmatic membrane. The inhibition of RNA and protein synthesis and of caspase activity also protected from cell death. This study shows that alterations in GSH levels change the neurotrophic effects of NO in midbrain cultures into neurotoxic. Under these conditions, NO triggers a programmed cell death with markers of both apoptosis and necrosis characterized by an early step of free radicals production followed by a late requirement for signalling on the sGC/cGMP/PKG pathway.     

Pockmarks enhance deep‐sea benthic biodiversity: a case study in the western Mediterranean Sea

Aim Pockmarks are craters on the sea floor formed by sub‐sea‐floor fluid expulsions, which occur world‐wide at all ocean depths. These habitats potentially host a highly specialized fauna that can exploit the hydrocarbons released. Pockmarks at relatively shallow depths can be easily destroyed by human activities, such as bottom trawling. In the present study, we investigated the combined effects of sea‐floor heterogeneity, rate of fluid emission and trophic conditions of different pockmarks on the biodiversity of the deep‐sea assemblages. Location Continental slope of the Gulf of Lions, western Mediterranean Sea, at water depths from 265 to 434 m. Methods We investigated the biodiversity associated with sea‐floor pockmarks that are both inactive and that have active gas emissions. Control sites were selected on the sea floor outside the influence of the gas seepage, both within and outside the pockmark fields. We examined the combined effects of: (i) sea‐floor heterogeneity; (ii) variable levels of fluid (gas) emissions; and (iii) trophic characteristics of the meiofaunal assemblage structure and nematode diversity. Results Sediments within the pockmark fields had lower meiofaunal abundance and biomass when compared with the surrounding sediments that were not influenced by the gas seepage. Although several higher taxa were absent in the pockmarks (e.g. Turbellaria, Tardigrada, Cumacea, Isopoda, Tanaidacea, Nemertina and Priapulida, which were present in the control areas), the richness of the nematode species within all of these pockmarks was very high. About 25% of the total species encountered in the deep‐sea sediments of the investigated areas was exclusively associated with these pockmarks. Main conclusions We conclude that both active and inactive pockmarks provide significant contributions to the regional (gamma) diversity of the continental slope in the western Mediterranean Sea, and thus the protection of these special and fragile habitats is highly relevant to the conservation of deep‐sea biodiversity.     

Role of the AAA protease Yme1 in folding of proteins in the intermembrane space of mitochondria

The vast majority of mitochondrial proteins are synthesized in the cytosol and transported into the organelle in a largely, if not completely, unfolded state. The proper function of mitochondria thus depends on folding of several hundreds of proteins in the various subcompartments of the organelle. Whereas folding of proteins in the mitochondrial matrix is supported by members of several chaperone families, very little is known about folding of proteins in the intermembrane space (IMS). We targeted dihydrofolate reductase (DHFR) as a model substrate to the IMS of yeast mitochondria and analyzed its folding. DHFR can fold in this compartment, and its aggregation upon heat shock can be prevented in an ATP-dependent manner. Yme1, an AAA (ATPases associated with diverse cellular activities) protease of the IMS, prevented aggregation of DHFR. Analysis of protein aggregates in mitochondria lacking Yme1 revealed the presence of a number of proteins involved in the establishment of mitochondrial ultrastructure, lipid metabolism, protein import, and respiratory growth. These findings explain the pleiotropic effects of deletion of YME1 and suggest an important role for Yme1 as a folding assistant, in addition to its proteolytic function, in the protein homeostasis of mitochondria     

Size diversity and species diversity relationships in fish assemblages of Western Palearctic lakes

Body size, coupled with abundance and taxonomy, may help to understand the mechanisms shaping community structure. Since the body size of fish is closely related to their trophic niche, size diversity (based on individual body size) of fish communities may capture intraspecific variations in fish trophic niches that are not detected by species diversity. Thus, the relationship between size diversity and species diversity may help to integrate variation at both intraspecific and interspecific levels. We studied the relationship between species diversity and size diversity as a measure of the degree of overlap in size among species and thereby the potential overlap in niches in a community. We hypothesized that the relationship between size diversity and species would be different across the European continent due to different levels of size overlap in fish communities. The data were derived from samplings of fish communities using standardised benthic gill nets in 363 lakes. At the continental scale, size diversity increased with species diversity; at the ecoregion scale, the slope of the relation changed across the continent, with the greatest mismatch occurring in northern Europe where communities comprised only one or a few species, but each of which exhibited a great range in size. There was an increase in slope towards the south with significant relations for four out of six ecoregions. The steeper size diversity‐species diversity slope at lower latitudes is attributable to a lower overlap in fish size and thus likely to finer niche separation. Our results also suggest that size diversity is not a strong surrogate for species diversity in European lake fish communities. Thus, particularly in fish communities composed of few species, measuring size diversity may help to detect potential functional variation which may be neglected by measuring species diversity alone.