Relief of hypoxia by angiogenesis promotes neural stem cell differentiation by targeting glycolysis

Blood vessels are part of the stem cell niche in the developing cerebral cortex, but their in vivo role in controlling the expansion and differentiation of neural stem cells (NSCs) in development has not been studied. Here, we report that relief of hypoxia in the developing cerebral cortex by ingrowth of blood vessels temporo‐spatially coincided with NSC differentiation. Selective perturbation of brain angiogenesis in vessel‐specific Gpr124 null embryos, which prevented the relief from hypoxia, increased NSC expansion at the expense of differentiation. Conversely, exposure to increased oxygen levels rescued NSC differentiation in Gpr124 null embryos and increased it further in WT embryos, suggesting that niche blood vessels regulate NSC differentiation at least in part by providing oxygen. Consistent herewith, hypoxia‐inducible factor (HIF)‐1α levels controlled the switch of NSC expansion to differentiation. Finally, we provide evidence that high glycolytic activity of NSCs is required to prevent their precocious differentiation in vivo. Thus, blood vessel function is required for efficient NSC differentiation in the developing cerebral cortex by providing oxygen and possibly regulating NSC metabolism.     

Characterization of heterotrophic growth and sesquiterpene production by Rhodobacter sphaeroides on a defined medium

Rhodobacter sphaeroides is a metabolically versatile bacterium capable of producing terpenes natively. Surprisingly, terpene biosynthesis in this species has always been investigated in complex media, with unknown compounds possibly acting as carbon and nitrogen sources. Here, a defined medium was adapted for R. sphaeroides dark heterotrophic growth, and was used to investigate the conversion of different organic substrates into the reporter terpene amorphadiene. The amorphadiene synthase was cloned in R. sphaeroides, allowing its biosynthesis via the native 2-methyl-d-erythritol-4-phosphate (MEP) pathway and, additionally, via a heterologous mevalonate one. The latter condition increased titers up to eightfold. Consequently, better yields and productivities to previously reported complex media cultivations were achieved. Productivity was further investigated under different cultivation conditions, including nitrogen and oxygen availability. This novel cultivation setup provided useful insight into the understanding of terpene biosynthesis in R. sphaeroides, allowing to better comprehend its dynamics and regulation during chemoheterotrophic cultivation.

     

High throughput measure of diversity in cytoplasmic and nuclear traits for unravelling geographic distribution of rosemary

In the present work, variability of both cytoplasmic and nuclear microsatellite traits was investigated with the aim of characterizing a set of rosemary germplasm resources (Salvia rosmarinus). Most of the materials were collected in Italy and France. High‐resolution melting curves were compared each other computing their Euclidean distances and estimating the differences within their principal component as a measure of genetic diversity. Mantel correlation results combined to linear discriminant analysis allowed examined populations to be divided in four principal groups corresponding to four geographic areas, with few interesting and discussed exceptions. As rosemary propagates by seeds coming from insect mediated pollination, steady wild populations can be expected to be in panmictic equilibrium. Gained results confirmed and extended precedent characterization of rosemary genotypes and are compatible with the distribution of other Mediterranean species, as well as with the presence of a glacial refugium in the north‐east area of Sardinia previously described. As the officinal use of this aromatic shrub is spreading, characterization and conservation of wild Mediterranean germplasm is gaining strategic importance. A core collection of 100 genotypes was pointed out as suitable for a cheaper biodiversity ex situ preservation as well as for subsequent metabolic and linkage disequilibrium analyses.     

Pesticide exposure affects flight dynamics and reduces flight endurance in bumblebees

The emergence of agricultural land use change creates a number of challenges that insect pollinators, such as eusocial bees, must overcome. Resultant fragmentation and loss of suitable foraging habitats, combined with pesticide exposure, may increase demands on foraging, specifically the ability to collect or reach sufficient resources under such stress. Understanding effects that pesticides have on flight performance is therefore vital if we are to assess colony success in these changing landscapes. Neonicotinoids are one of the most widely used classes of pesticide across the globe, and exposure to bees has been associated with reduced foraging efficiency and homing ability. One explanation for these effects could be that elements of flight are being affected, but apart from a couple of studies on the honeybee (Apis mellifera), this has scarcely been tested. Here, we used flight mills to investigate how exposure to a field realistic (10 ppb) acute dose of imidacloprid affected flight performance of a wild insect pollinator—the bumblebee, Bombus terrestris audax. Intriguingly, observations showed exposed workers flew at a significantly higher velocity over the first ¾ km of flight. This apparent hyperactivity, however, may have a cost because exposed workers showed reduced flight distance and duration to around a third of what control workers were capable of achieving. Given that bumblebees are central place foragers, impairment to flight endurance could translate to a decline in potential forage area, decreasing the abundance, diversity, and nutritional quality of available food, while potentially diminishing pollination service capabilities.     

Proteome analysis of whole saliva: a new tool for rheumatic diseases--the example of Sjögren's syndrome

Sj?gren's syndrome (pSS) is a systemic disease that affects salivary glands directly, and is therefore expected to influence the composition of human whole saliva (WS) fluid. The aim of this study was to characterize the WS proteins of pSS patients using a proteomic approach to assess a valid procedure to examine the global changes of the salivary protein profiles in connective tissue disorders. The WS proteins expressed in patients affected by pSS and healthy volunteers were analyzed using the 2-DE technique. The WS protein pattern was altered in pSS patients compared to controls, with a decrease in some of the typical salivary proteins. Particularly, a remarkable alteration of carbonic anhydrase VI was observed. Moreover, a comparison of WS protein profile of pSS patients with the one obtained from controls revealed a set of differentially expressed proteins. These proteins were related to acute and chronic inflammation while some others were involved in oxidative stress injury. These findings are in line with the systemic immuno-inflammatory aspects of pSS and open the possibility for a systematic search of diagnostic biomarkers and targets for therapeutic intervention in pSS.     

Mechanical response and conformational changes of alpha-actinin domains during unfolding: a molecular dynamics study

Alpha-actinin is a cytoskeleton-binding protein involved in the assembly and regulation of the actin filaments. In this work molecular dynamics method was applied to investigate the mechanical behaviour of the human skeletal muscle α-actinin. Five configurations were unfolded at an elongation speed of 0.1 nm/ps in order to investigate the conformational changes occurring during the extension process. Moreover, a sensitivity analysis at different velocities was performed for one of the R2–R3 spectrin-like repeat configuration extracted in order to evaluate the effect of the pulling speed on the mechanical behaviour of the molecule. Two different behaviours were recognized with respect to the pulling speed. In particular, at speed higher than 0.025 nm/ps a continuous rearrangement without evident force peaks was obtained, on the contrary at lower speed evident peaks in the range 500–750 pN were detected. R3 repeat resulted more stable than R2 during mechanical unfolding, due to the lower hydrophobic surface available to the solvent. The characterization of the R2–R3 units can be useful for the development of cytoskeleton network models based on stiffness values obtained by analyses performed at the molecular level.     

Strong genetic isolation despite wide distribution in a commercially exploited coastal shark

The common smoothhound, Mustelus mustelus, is an epibenthic species targeted by fisheries around the world driven by the increasing demand for shark products. Given the wide-spread occurrence of this species and corresponding lack of molecular data in many areas of said distribution, baseline molecular assessments of this commercially important shark may contribute to finer-scale analyses in areas in which this species is targeted. Therefore, population genetic analyses were conducted along the East Atlantic, from the Mediterranean Sea to the south-east coast of Africa, using microsatellite markers and the mitochondrial control region (mtCR). Overall, M. mustelus displayed low to moderate genetic diversity, with the Mediterranean populations appearing to exhibit the lowest mitochondrial diversity, and the west African populations displaying the lowest nuclear diversity. Microsatellite analysis indicated strong genetic differentiation between the three regions, with finer-scale population structure in each region, without correlation between genetic and geographical distance. For the mtCR sequences, a total of 18 haplotypes were identified, with a high degree of divergence discernable between the regions, largely in accordance with the microsatellite data. The study documents a remarkable level of population isolation across a vast area, suggesting little or no present-day connectivity among extant populations. The findings may serve as an essential baseline for global population management and commercial traceability of this threatened shark.

     

Lipid-Conjugated Rigidochromic Probe Discloses Membrane Alteration in Model Cells of Krabbe Disease

The plasma membrane of cells has a complex architecture based on the bidimensional liquid-crystalline bilayer arrangement of phospho- and sphingolipids, which in turn embeds several proteins and is connected to the cytoskeleton. Several studies highlight the spatial membrane organization into more ordered (L_o or lipid raft) and more disordered (L_d) domains. We here report on a fluorescent analog of the green fluorescent protein chromophore that, when conjugated to a phospholipid, enables the quantification of the L_o and L_d domains in living cells on account of its large fluorescence lifetime variation in the two phases. The domain composition is straightforwardly obtained by the phasor approach to confocal fluorescence lifetime imaging, a graphical method that does not require global fitting of the fluorescence decay in every spatial position of the sample. Our imaging strategy was applied to recover the domain composition in human oligodendrocytes at rest and under treatment with galactosylsphingosine (psychosine). Exogenous psychosine administration recapitulates many of the molecular fingerprints of a severe neurological disease, globoid cell leukodystrophy, better known as Krabbe disease. We found out that psychosine progressively destabilizes plasma membrane, as witnessed by a shrinking of the L_o fraction. The unchanged levels of galactosyl ceramidase, i.e., the enzyme lacking in Krabbe disease, upon psychosine treatment suggest that psychosine alters the plasma membrane structure by direct physical effect, as also recently demonstrated in model membranes.     

Environmental DNA metabarcoding as an effective and rapid tool for fish monitoring in canals

We focus on a case study along an English canal comparing environmental DNA (eDNA) metabarcoding with two types of electrofishing techniques (wade-and-reach and boom-boat). In addition to corroborating data obtained by electrofishing, eDNA provided a wider snapshot of fish assemblages. Given the semi-lotic nature of canals, we encourage the use of eDNA as a fast and cost-effective tool to detect and monitor whole fish communities.