Effects of climate-driven temperature changes on the diversity of freshwater macroinvertebrates

Increasing temperatures due to climate change were found to influence abundance and timing of species in numerous ways. Whereas many studies have investigated climate-induced effects on the phenology and abundance of single species, less is known about climate-driven shifts in the diversity and composition of entire communities. Analyses of long-term data sets provide the potential to reveal such relationships. We analysed time series of entire communities of macrozoobenthos in lakes and streams in Northern Europe. There were no direct linear effects of temperature and climate indices (North Atlantic Oscillation index) on species composition and diversity, but using multivariate statistics we were able to show that trends in average temperature have already had profound impacts on species composition in lakes. These significant temperature signals on species composition were evident even though we analysed comparatively short time periods of 10–15 years. Future climate shifts may thus induce strong variance in community composition. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users. Priority programme of the German Research Foundation—contribution 6.     

A gas-inducible expression system in HEK.EBNA cells applied to controlled proliferation studies by expression of p27(Kip1)

We describe an efficient inducible gene expression system in HEK.EBNA cells, a well-established cell system for the rapid transient expression of research-tool proteins. The transgene control system of choice is the novel acetaldehyde-inducible regulation (AIR) technology, which has been shown to modulate transgene levels following exposure of cells to acetaldehyde. For application in HEK.EBNA cells, AlcR transactivator plasmids were constructed and co-expressed with the secreted alkaline phosphatase (SEAP) gene under the control of a chimeric mammalian promoter (P(AIR)) for acetaldehyde-regulated expression. Several highly inducible transactivator cell lines were established. Adjustable transgene induction by gaseous acetaldehyde led to high induction levels and tight repression in transient expression trials and in stably transfected HEK.EBNA cell lines. Thus, the AIR technology can be used for inducible expression of any desired recombinant protein in HEK.EBNA cells. A possible application for inducible gene expression is a controlled proliferation strategy. Clonal HEK.EBNA cell lines, expressing the fungal transactivator protein AlcR, were engineered for gas-adjustable expression of the cell-cycle regulator p27(Kip1). We show that expression of p27(Kip1) via transient or stable transfection led to a G1-phase specific growth arrest of HEK.EBNA cells. Furthermore, production pools engineered for gas-adjustable expression of p27(Kip1) and constitutive expression of SEAP showed enhanced productive capacity.     

Fluorogenic surrogate substrates for toluene-degrading bacteria--are they useful for activity analysis?

Cultivated bacterial toluene degraders use one or several of four described pathways for the aerobic degradation of this priority groundwater contaminant. To be able to identify un-cultivated toluene-degrading bacteria within enriched or natural consortia, we attempted to develop a set of staining techniques that invariably label toluene-degrading bacteria while differentiating between the different degradation pathways. In the literature, we found suggestions for pathway-specific labels of individual cells that rely on the conversion of toluene surrogates into specific colored and fluorescent products. These surrogate substrates were phenylacetylene (PA), cinnamonitrile, 3-hydroxyphenylacetylene (3-HPA), and indole. We were able to confirm that the chromogenic reactions reliably verified the pathway-specific reactions of well-characterized toluene-degrading bacterial species. However, it was most surprising to find out that three (PA, 3-HPA and cinnamonitrile) of the four supplied surrogate substrates did not lead to any product fluorescence above the cultures' autofluorescence, neither inside of cells nor in supernatants. More disturbingly, the original surrogate compound 3-HPA was inherently fluorescent and found to stain cells at intensities that depended on their states in the cell cycle. Indoxyl originating from the surrogate substrate indole was the only fluorescent product that was formed. It was detected intracellularly when the cells were sealed with para-formaldehyde, but its appearance was unrelated to the presence of expressed toluene degradation pathways. These findings were scrutinized by fluorescence spectroscopy, fluorescence microscopy, and flow cytometry. Activity and growth of the test bacteria were determined by analyzing chromosome numbers and membrane integrity. Our results contradict literature reports that propose the surrogate fluorogenic substrates for the identification of toluene degraders and the identification of specific pathways used by them.     

Overexpression of Gremlin-1 in Patients with Loeys-Dietz Syndrome: Implications on Pathophysiology and Early Disease Detection

Backgrounds

The Loeys-Dietz syndrome (LDS) is an inherited connective tissue disorder caused by mutations in the transforming growth factor β (TGF-β) receptors TGFBR1 or TGFBR2. Most patients with LDS develop severe aortic aneurysms resulting in early need of surgical intervention. In order to gain further insight into the pathophysiology of the disorder, we investigated circulating outgrowth endothelial cells (OEC) from the peripheral blood of LDS patients from a cohort of 23 patients including 6 patients with novel TGF-β receptor mutations.

Methods and Results

We performed gene expression profiling of OECs using microarray analysis followed by quantitative PCR for verification of gene expression. Compared to OECs of age- and sex-matched healthy controls, OECs isolated from three LDS patients displayed altered expression of several genes belonging to the TGF-β pathway, especially those affecting bone morphogenic protein (BMP) signalling including BMP2, BMP4 and BMPR1A. Gene expression of BMP antagonist Gremlin-1 (GREM1) showed the most prominent up-regulation. This increase was confirmed at the protein level by immunoblotting of LDS-OECs. In immunohistochemistry, abundant Gremlin-1 protein expression could be verified in endothelial cells as well as smooth muscle cells within the arterial media. Furthermore, Gremlin-1 plasma levels of LDS patients were significantly elevated compared to healthy control subjects.

Conclusions

These findings open new avenues in the understanding of the pathogenesis of Loeys-Dietz syndrome and the development of new diagnostic serological methods for early disease detection.     

Genetic dissection of temperature-dependent sorghum growth during juvenile development

Key message

Promising genome regions for improving cold tolerance of sorghum were identified on chromosomes SBI-01, SBI-03, SBI-07, and SBI-10. Chlorophyll fluorescence had no major effect on growth rates at low temperatures.

Abstract

Developing fast growing sorghum seedlings is an important breeding goal for temperate climates since low springtime temperatures are resulting in a prolonged juvenile development. The adaptation of sorghum to tropical and subtropical highlands gives hint for certain genetic variation. The goals of the present study were to detect marker-trait associations for leaf and dry matter growth rate and for chlorophyll fluorescence and content (SPAD) in relation to temperature. A diversity set comprising 194 genotypes was tested in eight controlled environments with temperatures ranging from 9.4 to 20.8 °C. Significant marker-trait associations (p < 0.05) were identified for each individual temperature regime and on the parameters of regression analyses describing the responses of growth or chlorophyll related traits to temperatures. The diversity set was fingerprinted with 171 diversity array technology (DArT) and 31 simple-sequence repeat (SSR) markers. SSRs were used to analyze the population structure while association studies were performed on DArT markers. Promising marker-trait associations for growth rates in relation to temperature were detected on chromosomes SBI-01, SBI-03, SBI-07, and SBI-10. Many promising loci were also significantly associated to the results obtained in individual low-temperature environments. Marker-trait associations for chlorophyll content and fluorescence did occasionally co-locate to those for growth during juvenile development but there was no evidence supporting our hypothesis that seedling growth at low temperatures is largely influenced by SPAD or fluorescence.     

DNA staining in agarose gels with Zn²+-cyclen-pyrene

A pyrene-labeled Zn2+-cyclen complex for the staining of DNA in agarose gels is reported. The metal chelate coordinates reversibly to the DNA phosphate backbone, which induces the formation of pyrene excimers. The typical pyrene excimer emission is used for the detection of the DNA. Staining is limited to agarose gels and is less sensitive than ethidium bromide, but DNA amounts as low as 10 ng and short DNA strands (～300 b.p.) are detectable. Gel extraction as a standard technique in molecular biology was successfully performed after staining with Zn2+-cyclen-pyrene. Cytotoxicity tests on HeLa and V-79 cells reveal that the zinc-cyclen pyrene probe is significant less toxic compared to ethidium bromide.     

Predation on mutualists can reduce the strength of trophic cascades

Ecologists have put forth several mechanisms to predict the strength of predator effects on producers (a trophic cascade). We suggest a novel mechanism – in systems in which mutualists of plants are present and important, predators can have indirect negative effects on producers through their consumption of mutualists. The strength of predator effects on producers will depend on their relative consumption of mutualists and antagonists, and on the relative importance of each to producer population dynamics. In a meta-analysis of experiments that examine the effects of predator reduction on the pollination and reproductive success of plants, we found that the indirect negative effects of predators on plants are quite strong. Most predator removal experiments measure the strength of predator effects on producers through the antagonist pathway; we suggest that a more complete understanding of the role of predators will be achieved by simultaneously considering the effects of predators on plant mutualists.