Losing ground: past history and future fate of Arctic small mammals in a changing climate

According to the IPCC, the global average temperature is likely to increase by 1.4–5.8 °C over the period from 1990 to 2100. In Polar regions, the magnitude of such climatic changes is even larger than in temperate and tropical biomes. This amplified response is particularly worrisome given that the so‐far moderate warming is already impacting Arctic ecosystems. Predicting species responses to rapid warming in the near future can be informed by investigating past responses, as, like the rest of the planet, the Arctic experienced recurrent cycles of temperature increase and decrease (glacial–interglacial changes) in the past. In this study, we compare the response of two important prey species of the Arctic ecosystem, the collared lemming and the narrow‐skulled vole, to Late Quaternary climate change. Using ancient DNA and Ecological Niche Modeling (ENM), we show that the two species, which occupy similar, but not identical ecological niches, show markedly different responses to climatic and environmental changes within broadly similar habitats. We empirically demonstrate, utilizing coalescent model‐testing approaches, that collared lemming populations decreased substantially after the Last Glacial Maximum; a result consistent with distributional loss over the same period based on ENM results. Given this strong association, we projected the current niche onto future climate conditions based on IPCC 4.0 scenarios, and forecast accelerating loss of habitat along southern range boundaries with likely associated demographic consequences. Narrow‐skulled vole distribution and demography, by contrast, was only moderately impacted by past climatic changes, but predicted future changes may begin to affect their current western range boundaries. Our work, founded on multiple lines of evidence suggests a future of rapidly geographically shifting Arctic small mammal prey communities, some of whom are on the edge of existence, and whose fate may have ramifications for the whole Arctic food web and ecosystem.     

Neuroprotection against traumatic brain injury by a peptide derived from the collapsin response mediator protein 2 (CRMP2)

Neurological disabilities following traumatic brain injury (TBI) may be due to excitotoxic neuronal loss. The excitotoxic loss of neurons following TBI occurs largely due to hyperactivation of N-methyl-d-aspartate receptors (NMDARs), leading to toxic levels of intracellular Ca(2+). The axon guidance and outgrowth protein collapsin response mediator protein 2 (CRMP2) has been linked to NMDAR trafficking and may be involved in neuronal survival following excitotoxicity. Lentivirus-mediated CRMP2 knockdown or treatment with a CRMP2 peptide fused to HIV TAT protein (TAT-CBD3) blocked neuronal death following glutamate exposure probably via blunting toxicity from delayed calcium deregulation. Application of TAT-CBD3 attenuated postsynaptic NMDAR-mediated currents in cortical slices. In exploring modulation of NMDARs by TAT-CBD3, we found that TAT-CBD3 induced NR2B internalization in dendritic spines without altering somal NR2B surface expression. Furthermore, TAT-CBD3 reduced NMDA-mediated Ca(2+) influx and currents in cultured neurons. Systemic administration of TAT-CBD3 following a controlled cortical impact model of TBI decreased hippocampal neuronal death. These findings support TAT-CBD3 as a novel neuroprotective agent that may increase neuronal survival following injury by reducing surface expression of dendritic NR2B receptors.     

Collapsin Response Mediator Protein 2 (CRMP2) Interacts with N-Methyl-d-aspartate (NMDA) Receptor and Na+/Ca2+ Exchanger and Regulates Their Functional Activity

Collapsin response mediator protein 2 (CRMP2) is traditionally viewed as an axonal growth protein involved in axon/dendrite specification. Here, we describe novel functions of CRMP2. A 15-amino acid peptide from CRMP2, fused to the TAT cell-penetrating motif of the HIV-1 protein, TAT-CBD3, but not CBD3 without TAT, attenuated N-methyl-d-aspartate receptor (NMDAR) activity and protected neurons against glutamate-induced Ca²⁺ dysregulation, suggesting the key contribution of CRMP2 in these processes. In addition, TAT-CBD3, but not CBD3 without TAT or TAT-scramble peptide, inhibited increases in cytosolic Ca²⁺ mediated by the plasmalemmal Na⁺/Ca²⁺ exchanger (NCX) operating in the reverse mode. Co-immunoprecipitation experiments revealed an interaction between CRMP2 and NMDAR as well as NCX3 but not NCX1. TAT-CBD3 disrupted CRMP2-NMDAR interaction without change in NMDAR localization. In contrast, TAT-CBD3 augmented the CRMP2-NCX3 co-immunoprecipitation, indicating increased interaction or stabilization of a complex between these proteins. Immunostaining with an anti-NCX3 antibody revealed that TAT-CBD3 induced NCX3 internalization, suggesting that both reverse and forward modes of NCX might be affected. Indeed, the forward mode of NCX, evaluated in experiments with ionomycin-induced Ca²⁺ influx into neurons, was strongly suppressed by TAT-CBD3. Knockdown of CRMP2 with short interfering RNA (siRNA) prevented NCX3 internalization in response to TAT-CBD3 exposure. Moreover, CRMP2 down-regulation strongly attenuated TAT-CBD3-induced inhibition of reverse NCX. Overall, our results demonstrate that CRMP2 interacts with NCX and NMDAR and that TAT-CBD3 protects against glutamate-induced Ca²⁺ dysregulation most likely via suppression of both NMDAR and NCX activities. Our results further clarify the mechanism of action of TAT-CBD3 and identify a novel regulatory checkpoint for NMDAR and NCX function based on CRMP2 interaction with these proteins.     

Genetic diversity and genetic structure of the Siberian roe deer (<Emphasis Type="Italic">Capreolus pygargus</Emphasis>) populations from Asia

Background

The roe deer, Capreolus sp., is one of the most widespread meso-mammals of Palearctic distribution, and includes two species, the European roe deer, C. capreolus inhabiting mainly Europe, and the Siberian roe deer, C. pygargus, distributed throughout continental Asia. Although there are a number of genetic studies concerning European roe deer, the Siberian roe deer has been studied less, and none of these studies use microsatellite markers. Natural processes have led to genetic structuring in wild populations. To understand how these factors have affected genetic structure and connectivity of Siberian roe deer, we investigated variability at 12 microsatellite loci for Siberian roe deer from ten localities in Asia.

Results

Moderate levels of genetic diversity (H _E = 0.522 to 0.628) were found in all populations except in Jeju Island, South Korea, where the diversity was lowest?(H _E?= 0.386). Western populations showed relatively low genetic diversity and higher degrees of genetic differentiation compared with eastern populations (mean Ar = 3.54 (east), 2.81 (west), mean F _ST = 0.122). Bayesian-based clustering analysis revealed the existence of three genetically distinct groups (clusters) for Siberian roe deer, which comprise of the Southeastern group (Mainland Korea, Russian Far East, Trans-Baikal region and Northern part of Mongolia), Northwestern group (Western Siberia and Ural in Russia) and Jeju Island population. Genetic analyses including AMOVA (F _RT = 0.200), Barrier and PCA also supported genetic differentiation among regions separated primarily by major mountain ridges, suggesting that mountains played a role in the genetic differentiation of Siberian roe deer. On the other hand, genetic evidence also suggests an ongoing migration that may facilitate genetic admixture at the border areas between two groups.

Conclusions

Our results reveal an apparent pattern of genetic differentiation among populations inhabiting Asia, showing moderate levels of genetic diversity with an east-west gradient. The results suggest at least three distinct management units of roe deer in continental Asia, although genetic admixture is evident in some border areas. The insights obtained from this study shed light on management of Siberian roe deer in Asia and may be applied in conservation of local populations of Siberian roe deer.

     

Calcium-dependent spontaneously reversible remodeling of brain mitochondria

An exposure of cultured hippocampal neurons expressing mitochondrially targeted enhanced yellow fluorescent protein to excitotoxic glutamate resulted in reversible mitochondrial remodeling that in many instances could be interpreted as swelling. Remodeling was not evident if glutamate receptors were blocked with MK801, if Ca(2+) was omitted or substituted for Sr(2+) in the bath solution, if neurons were treated with carbonylcyanide p-trifluoromethoxyphenylhydrazone to depolarize mitochondria, or if neurons were pretreated with cyclosporin A or N-methyl-4-isoleucine-cyclosporin (NIM811) to inhibit the mitochondrial permeability transition. In the experiments with isolated brain synaptic or nonsynaptic mitochondria, Ca(2+) triggered transient, spontaneously reversible cyclosporin A-sensitive swelling closely resembling remodeling of organelles in cultured neurons. The swelling was accompanied by the release of cytochrome c, Smac/DIABLO, Omi/HtrA2, and AIF but not endonuclease G. Depolarization with carbonylcyanide p-trifluoromethoxyphenylhydrazone or inhibition of the Ca(2+) uniporter with Ru360 prevented rapid onset of the swelling. Sr(2+) depolarized mitochondria but failed to induce swelling. Neither inhibitors of the large conductance Ca(2+)-activated K(+) channel (charybdotoxin, iberiotoxin, quinine, and Ba(2+)) nor inhibitors of the mitochondrial ATP-sensitive K(+) channel (5-hydroxydecanoate and glibenclamide) suppressed swelling. Quinine, dicyclohexylcarbodiimide, and Mg(2+), inhibitors of the mitochondrial K(+)/H(+) exchanger, as well as external alkalization inhibited a recovery phase of the reversible swelling. In contrast to brain mitochondria, liver and heart mitochondria challenged with Ca(2+) experienced sustained swelling without spontaneous recovery. The proposed model suggests an involvement of the Ca(2+)-dependent transient K(+) influx into the matrix causing mitochondrial swelling followed by activation of the K(+)/H(+) exchanger leading to spontaneous mitochondrial contraction both in situ and in vitro.     

Production of Brassica olereceae (+Arabidopsis thaliana) and Brassica napus cell lines resistant to spectinomycin/streptomycin as a result of plastome genetic transformation

Plastid genetic transformation has been performed using both the PEG-treatment of protoplasts of somatic hybrids of B. oleracea carrying A. thaliana chloroplasts and the particle bombardment of regenerable calluses of B. napus sv. Westar. The chloroplast transformation vector pCB040 carried resistance (aadA) gene flanked by rapeseed plastid DNA sequences to target its insertion between the trnV-rps7 fragments. Selection of transplastomic cell lines has been performed according to their ability to grow on the medium supplied with spectinomycin and streptomycin in high concentrations. Antibiotic resistant cell lines have been obtained using the both transformation methods. The presence of the aadA gene in the A. thaliana and B. napus plastomes was confirmed by PCR analysis for two cell lines of B. oleracea (+ A. thaliana) and three lines of B. napus.     

The spring energy budget of the algal mat community in a Crimean hypersaline lake determined by microcalorimetry

The energy contents (standing stock) of the floating mat formed by the green alga Cladophora sivaschensis and the energy transfers through it were quantified for a shallow hypersaline lake (at Cape Khersones, Crimea, Ukraine) during the spring months. Appropriate direct calorimetric techniques were applied to: (i) measure the heat energy dissipated by the mat community and by the free bacterioplankton in the water column below it; and (ii) differentiate between the heat flows by the heterotrophic and the phototrophic components of the community. It was shown that Cladophora biomass reached a peak of 579.5 g C m^–2, contributing more than 99.6% of the total mat community. Throughout the spring, the total bacterial energy transfer (6 to 23 mW m^–2) was as little as 1.1 to 2.6% of the total heat dissipated by the microplankton community. The rest of the estimated heat energy (584 to 1488 mW m^–2) was associated with Cladophora metabolism. In the spring community: (i) the rate of biomass accumulation in the lake photic layer significantly exceeded its heterotrophic mineralisation; (ii) the efficiency of the microbial loop was too low to process even a minor part of the accumulated organic matter. The microcalorimetric technique was shown to be a highly promising approach for further studies of natural microbial mats and biofilms, biological systems with complex metabolism that involves not only aerobic processes but also anaerobic catabolism under local hypoxic/microxic conditions.     

Structural analysis of the heterodimeric type IIS restriction endonuclease R.BspD6I acting as a complex between a monomeric site-specific nickase and a catalytic subunit

The heterodimeric restriction endonuclease R.BspD6I from Bacillus species D6 recognizes a pseudosymmetric sequence and cuts both DNA strands outside the recognition sequence. The large subunit, Nt.BspD6I, acts as a type IIS site-specific monomeric nicking endonuclease. The isolated small subunit, ss.BspD6I, does not bind DNA and is not catalytically active. We solved the crystal structures of Nt.BspD6I and ss.BspD6I at high resolution. Nt.BspD6I consists of three domains, two of which exhibit structural similarity to the recognition and cleavage domains of FokI. ss.BspD6I has a fold similar to that of the cleavage domain of Nt.BspD6I, each containing a PD-(D/E)XK motif and a histidine as an additional putative catalytic residue. In contrast to the DNA-bound FokI structure, in which the cleavage domain is rotated away from the DNA, the crystal structure of Nt.BspD6I shows the recognition and cleavage domains in favorable orientations for interactions with DNA. Docking models of complexes of Nt.BspD6I and R.BspD6I with cognate DNA were constructed on the basis of structural similarity to individual domains of FokI, R.BpuJI and HindIII. A three-helix bundle forming an interdomain linker in Nt.BspD6I acts as a rigid spacer adjusting the orientations of the spatially separated domains to match the distance between the recognition and cleavage sites accurately.     

Obtaining of intertribal Brassica juncea+Arabidopsis thaliana somatic hybrids and study of transgenic trait behaviour

Intertribal somatic hybridization between wild type Brassica juncea (L.) Czern. & Coss. and transgenic A. thaliana L. has been carried out. Genome of A. thaliana plants contained heterologous transposable element Spm/dSpm, reporter GUS gene, selective genes for kanamycin- (npt II) and phosphinothricin (bar) resistance. Hybrid nature of obtained plants was confirmed with their morphology, GUS hystochemical assay, PCR-RFLP, RAPD and isozyme analyses. It was determined that heterologous transposable element Spm/dSpm is able to function in hybrid plants. There was no complete elimination of A. thaliana genetic material in the hybrids and the transgenes were stably maintained.     

Production of transgenic sugarbeet (Beta vulgaris L.) plants of O-type using Agrobacterium tumefaciens

A method of Agrobacterium-mediated genetic transformation of sugarbeet (Beta vulgaris L.) by vacuum infiltration has been developed. Transgenic sugarbeet plants of Ukraine breeding were selected for their resistance either to the antibiotic kanamycin or to the herbicide glufosinate ammonium. Integration of transgenes was confirmed by PCR and GUS-assay.