Activation of PDGF-CC by tissue plasminogen activator impairs blood-brain barrier integrity during ischemic stroke

Thrombolytic treatment of ischemic stroke with tissue plasminogen activator (tPA) is markedly limited owing to concerns about hemorrhagic complications and the requirement that tPA be administered within 3 h of symptoms. Here we report that tPA activation of latent platelet-derived growth factor-CC (PDGF-CC) may explain these limitations. Intraventricular injection of tPA or active PDGF-CC, in the absence of ischemia, leads to significant increases in cerebrovascular permeability. In contrast, co-injection of neutralizing antibodies to PDGF-CC with tPA blocks this increased permeability, indicating that PDGF-CC is a downstream substrate of tPA within the neurovascular unit. These effects are mediated through activation of PDGF-alpha receptors (PDGFR-alpha) on perivascular astrocytes, and treatment of mice with the PDGFR-alpha antagonist imatinib after ischemic stroke reduces both cerebrovascular permeability and hemorrhagic complications associated with late administration of thrombolytic tPA. These data demonstrate that PDGF signaling regulates blood-brain barrier permeability and suggest potential new strategies for stroke treatment.     

Dissecting sensor functions in cell wall integrity signaling in Kluyveromyces lactis.

KlWSC1, KlWSC2/3 and KlMID2, which encode putative plasma membrane sensors for cell wall integrity signaling in Kluyveromyces lactis, were cloned and characterized. Double and triple deletion mutants show severe cell integrity defects, indicating overlapping functions. The Klwsc1 Klmid2 double deletion phenotype can be suppressed by overexpression of the downstream components KlROM2, KlPKC1 and KlBCK1. KlWsc1 sensor domain analyses showed that an amino-terminal elongation as well as an extension within the cytoplasmic domain are dispensable for function. Heterologous complementation by KlMID2 and KlWSC1 in Saccharomyces cerevisiae is only achieved upon overexpression. In contrast to ScMID2, ScWSC1 complements in K. lactis. Functional studies with chimeric Mid2 constructs indicate that species specificity is mainly conferred by the extracellular domain. Sensor-GFP fusions localize to the plasma membrane, with a cell cycle dependent distribution of KlWsc1-GFP. Both Wsc-type sensors concentrate in discrete spots within the plasma membrane.     

Evaluation of Epicoccum nigrum for growth, morphology and production of natural colorants in liquid media and on a solid rice medium

Four nonpathogenic and nontoxigenic Epicoccum nigrum strains were evaluated for their growth, morphology and pigment producing ability in three complex and one defined liquid media. Epicoccum nigrum IBT 41028 produced pigments in all the four media tested with a maximum pigment of 3.68 AU at 410 nm in M1 medium (unoptimized) containing 5 g/l yeast autolysate. The color hue of the crude pigment extracts ranged from 74 to 102 exhibiting dark orange to green-yellow color. Pelleted morphology was shown to have a positive influence on the pigment production by E. nigrum strain IBT 41028 in the liquid media, and the use of Bis-tris buffer was found to diminish or reduce the pellet formation. Since Monascus is a well known pigment producer on rice. Pigment producing ability of E. nigrum IBT 41028 was tested on rice and compared to liquid media with Monascus ruber IBT 7904 as control. Though, both genera preferred rice but E. nigrum produced 4.6 folds higher pigment in the liquid unoptimized fermentation medium compared to M. ruber. Solid phase extraction and subsequently HPLC-DAD analysis of the crude pigment extracts showed qualitative as well as quantitative variation in the pigment composition under solid and liquid cultivations.     

Binomial mitotic segregation of MYCN-carrying double minutes in neuroblastoma illustrates the role of randomness in oncogene amplification

Background

Amplification of the oncogene MYCN in double minutes (DMs) is a common finding in neuroblastoma (NB). Because DMs lack centromeric sequences it has been unclear how NB cells retain and amplify extrachromosomal MYCN copies during tumour development.

Principal Findings

We show that MYCN-carrying DMs in NB cells translocate from the nuclear interior to the periphery of the condensing chromatin at transition from interphase to prophase and are preferentially located adjacent to the telomere repeat sequences of the chromosomes throughout cell division. However, DM segregation was not affected by disruption of the telosome nucleoprotein complex and DMs readily migrated from human to murine chromatin in human/mouse cell hybrids, indicating that they do not bind to specific positional elements in human chromosomes. Scoring DM copy-numbers in ana/telophase cells revealed that DM segregation could be closely approximated by a binomial random distribution. Colony-forming assay demonstrated a strong growth-advantage for NB cells with high DM (MYCN) copy-numbers, compared to NB cells with lower copy-numbers. In fact, the overall distribution of DMs in growing NB cell populations could be readily reproduced by a mathematical model assuming binomial segregation at cell division combined with a proliferative advantage for cells with high DM copy-numbers.

Conclusion

Binomial segregation at cell division explains the high degree of MYCN copy-number variability in NB. Our findings also provide a proof-of-principle for oncogene amplification through creation of genetic diversity by random events followed by Darwinian selection.     

Screening of novel bacteria for the 2,3-butanediol production

Biotechnologically produced 2,3-butanediol (2,3-BDO) is a potential starting material for industrial bulk chemicals such as butadiene or methyl ethyl ketone which are currently produced from fossil feedstocks. So far, the highest 2,3-BDO concentrations have been obtained with risk group 2 microorganisms. In this study, three risk group 1 microorganisms are presented that are so far unknown for an efficient production of 2,3-BDO. The strains Bacillus atrophaeus NRS-213, Bacillus mojavensis B-14698, and Bacillus vallismortis B-14891 were evaluated regarding their ability to produce high 2,3-BDO concentrations with a broad range of different carbon sources. A maximum 2,3-BDO concentration of 60.4 g/L was reached with the strain B. vallismortis B-14891 with an initial glucose concentration of 200 g/L within 55 h in a batch cultivation. Besides glucose, B. vallismortis B-14891 converts 14 different substrates that can be obtained from residual biomass sources to 2,3-BDO. Therefore B. vallismortis B-14891 is a promising candidate for the large-scale production of 2,3-BDO with low-cost substrates.

     

Silver fir and Douglas fir are more tolerant to extreme droughts than Norway spruce in south‐western Germany

Improving our understanding of the potential of forest adaptation is an urgent task in the light of predicted climate change. Long‐term alternatives for susceptible yet economically important tree species such as Norway spruce (Picea abies) are required, if the frequency and intensity of summer droughts will continue to increase. Although Silver fir (Abies alba) and Douglas fir (Pseudotsuga menziesii) have both been described as drought‐tolerant species, our understanding of their growth responses to drought extremes is still limited. Here, we use a dendroecological approach to assess the resistance, resilience, and recovery of these important central Europe to conifer species the exceptional droughts in 1976 and 2003. A total of 270 trees per species were sampled in 18 managed mixed‐species stands along an altitudinal gradient (400–1200 m a.s.l.) at the western slopes of the southern and central Black Forest in southwest Germany. While radial growth in all species responded similarly to the 1976 drought, Norway spruce was least resistant and resilient to the 2003 summer drought. Silver fir showed the overall highest resistance to drought, similarly to Douglas fir, which exhibited the widest growth rings. Silver fir trees from lower elevations were more drought prone than trees at higher elevations. Douglas fir and Norway spruce, however, revealed lower drought resilience at higher altitudes. Although the 1976 and 2003 drought extremes were quite different, Douglas fir maintained consistently the highest radial growth. Although our study did not examine population‐level responses, it clearly indicates that Silver fir and Douglas fir are generally more resistant and resilient to previous drought extremes and are therefore suitable alternatives to Norway spruce; Silver fir more so at higher altitudes. Cultivating these species instead of Norway spruce will contribute to maintaining a high level of productivity across many Central European mountain forests under future climate change.     

Greater temperature sensitivity of plant phenology at colder sites: implications for convergence across northern latitudes

Warmer temperatures are accelerating the phenology of organisms around the world. Temperature sensitivity of phenology might be greater in colder, higher latitude sites than in warmer regions, in part because small changes in temperature constitute greater relative changes in thermal balance at colder sites. To test this hypothesis, we examined up to 20 years of phenology data for 47 tundra plant species at 18 high‐latitude sites along a climatic gradient. Across all species, the timing of leaf emergence and flowering was more sensitive to a given increase in summer temperature at colder than warmer high‐latitude locations. A similar pattern was seen over time for the flowering phenology of a widespread species, Cassiope tetragona. These are among the first results highlighting differential phenological responses of plants across a climatic gradient and suggest the possibility of convergence in flowering times and therefore an increase in gene flow across latitudes as the climate warms.     

Effects of ocean acidification increase embryonic sensitivity to thermal extremes in Atlantic cod,Gadus morhua

Thermal tolerance windows serve as a powerful tool for estimating the vulnerability of marine species and their life stages to increasing temperature means and extremes. However, it remains uncertain to which extent additional drivers, such as ocean acidification, modify organismal responses to temperature. This study investigated the effects of CO₂‐driven ocean acidification on embryonic thermal sensitivity and performance in Atlantic cod, Gadus morhua, from the Kattegat. Fertilized eggs were exposed to factorial combinations of two PCO₂ conditions (400 μatm vs. 1100 μatm) and five temperature treatments (0, 3, 6, 9 and 12 °C), which allow identifying both lower and upper thermal tolerance thresholds. We quantified hatching success, oxygen consumption (MO₂) and mitochondrial functioning of embryos as well as larval morphometrics at hatch and the abundance of acid–base‐relevant ionocytes on the yolk sac epithelium of newly hatched larvae. Hatching success was high under ambient spawning conditions (3–6 °C), but decreased towards both cold and warm temperature extremes. Elevated PCO₂ caused a significant decrease in hatching success, particularly at cold (3 and 0 °C) and warm (12 °C) temperatures. Warming imposed limitations to MO₂ and mitochondrial capacities. Elevated PCO₂ stimulated MO₂ at cold and intermediate temperatures, but exacerbated warming‐induced constraints on MO₂, indicating a synergistic interaction with temperature. Mitochondrial functioning was not affected by PCO₂. Increased MO₂ in response to elevated PCO₂ was paralleled by reduced larval size at hatch. Finally, ionocyte abundance decreased with increasing temperature, but did not differ between PCO₂ treatments. Our results demonstrate increased thermal sensitivity of cod embryos under future PCO₂ conditions and suggest that acclimation to elevated PCO₂ requires reallocation of limited resources at the expense of embryonic growth. We conclude that ocean acidification constrains the thermal performance window of embryos, which has important implication for the susceptibility of cod to projected climate change.     

Phenology and temperature‐dependent development of Ceutorhynchus assimilis,a potential biological control agent for Lepidium draba

Lepidium draba (Brassicaceae) is a major concern for agriculture and biodiversity in the western United States. As current control methods do not provide long‐term, sustainable solutions, research has been conducted to find biological control agents. Ceutorhynchus assimilis is one of the currently investigated candidates. Known as oligophagous in the literature, a specialist clade of this root‐galling weevil exists in southern Europe. This raised the question of its ability to survive in colder climates in the target range. We investigated the phenology of C. assimilis in the field in southern France (specialist clade) and Romania (generalist clade) and measured various temperature‐dependent parameters in the laboratory. In both ranges, weevils were univoltine. Oviposition in autumn started later in France compared to Romania, while mature larvae exited galls (to pupate in the soil) earlier the following year. On average, 25% and 32% of galls from France and Romania were completely below the soil surface, respectively, and this appeared to depend on soil substrate. Weevils transported from France to Romania were able to develop, but at a much lower rate than Romanian weevils. Mortality of overwintering larvae of both clades increased with decreasing temperature and exposure time. At ?5°C, lethal times Lt₅₀ and Lt₉₅ were 15 and 42 days for the specialist clade and 26 and 72 days for the generalist clade. A higher proportion of third instar larvae compared to first and second instar larvae survived. Pupation time at different temperatures did not differ between weevils from France or Romania. A climate match model (comparing winter temperatures) indicated that the specialist clade of C. assimilis from France has the potential to establish in some parts of the target range (e.g. Washington, Oregon, California). However, temperature extremes and winters without snow cover will likely limit its establishment unless rapid adaptive evolution takes place.