A method for in situ monitoring of the isotope composition of tree xylem water using laser spectroscopy

Field studies analyzing the stable isotope composition of xylem water are providing important information on ecosystem water relations. However, the capacity of stable isotopes to characterize the functioning of plants in their environment has not been fully explored because of methodological constraints on the extent and resolution at which samples could be collected and analysed. Here, we introduce an in situ method offering the potential to continuously monitor the stable isotope composition of tree xylem water via its vapour phase using a commercial laser‐based isotope analyser and compact microporous probes installed into the xylem. Our technique enables efficient high‐frequency measurement with intervals of only a few minutes per sample while eliminating the need for costly and cumbersome destructive collection of plant material and laboratory‐based processing. We present field observations of xylem water hydrogen and oxygen isotope compositions obtained over several days including a labelled irrigation event and compare them against results from concurrent destructive sampling with cryogenic distillation and mass spectrometric analysis. The data demonstrate that temporal changes as well as spatial patterns of integration in xylem water isotope composition can be resolved through direct measurement. The new technique can therefore present a valuable tool to study the hydraulic architecture and water utilization of trees.     

Treatment of poor-risk myelodysplastic syndromes and acute myeloid leukemia with a combination of 5-azacytidine and valproic acid

5-azacytidine (AZA) has become standard treatment for patients with higher-risk myelodysplastic syndrome (MDS). Response rate is about 50% and response duration is limited. Histone deactylase (HDAC) inhibitors are attractive partners for epigenetic combination therapy. We treated 24 patients with AZA (100?mg/m(2), 5?days) plus valproate (VPA; continuous dosing, trough serum level 80-110?μg/ml). According to WHO classification, 5 patients had MDS, 2 had MDS/MPD, and 17 had acute myeloid leukemia (AML). Seven patients (29%) had previously received intensive chemotherapy, and five had previous HDAC inhibitor treatment. The overall response rate was 37% in the entire cohort but significantly higher (57%) in previously untreated patients, especially those with MDS (64%). Seven (29%) patients achieved CR (29%) and two PR (8%), respectively. Hematological CR was accompanied by complete cytogenetic remission according to conventional cytogenetics in all evaluable cases. Some patients also showed complete remission according to FISH on bone marrow mononuclear cells and CD34(+) peripheral blood cells, as well as by follow-up of somatic mitochondrial DNA mutations. Four additional patients achieved at least marrow remissions. Factors influencing response were AML (vs. MDS), marrow blast count, pretreatment, transfusion dependency, concomitant medication with hydroxyurea, and valproic acid (VPA) serum level. This trial is the first to assess the combination of AZA plus VPA without additional ATRA. A comparatively good CR rate, relatively short time to response, and the influence of VPA serum levels on response suggest that VPA provided substantial additional benefit. However, the importance of HDAC inhibitors in epigenetic combination therapy can only be proven by randomized trials.     

A -436C>A polymorphism in the human FAS gene promoter associated with severe childhood malaria

Human genetics and immune responses are considered to critically influence the outcome of malaria infections including life-threatening syndromes caused by Plasmodium falciparum. An important role in immune regulation is assigned to the apoptosis-signaling cell surface receptor CD95 (Fas, APO-1), encoded by the gene FAS. Here, a candidate-gene association study including variant discovery at the FAS gene locus was carried out in a case-control group comprising 1,195 pediatric cases of severe falciparum malaria and 769 unaffected controls from a region highly endemic for malaria in Ghana, West Africa. We found the A allele of c.−436C>A (rs9658676) located in the promoter region of FAS to be significantly associated with protection from severe childhood malaria (odds ratio 0.71, 95% confidence interval 0.58–0.88, p_empirical = 0.02) and confirmed this finding in a replication group of 1,412 additional severe malaria cases and 2,659 community controls from the same geographic area. The combined analysis resulted in an odds ratio of 0.71 (95% confidence interval 0.62–0.80, p = 1.8×10⁻⁷, n = 6035). The association applied to c.−436AA homozygotes (odds ratio 0.47, 95% confidence interval 0.36–0.60) and to a lesser extent to c.−436AC heterozygotes (odds ratio 0.73, 95% confidence interval 0.63–0.84), and also to all phenotypic subgroups studied, including severe malaria anemia, cerebral malaria, and other malaria complications. Quantitative FACS analyses assessing CD95 surface expression of peripheral blood mononuclear cells of naïve donors showed a significantly higher proportion of CD69⁺CD95⁺ cells among persons homozygous for the protective A allele compared to AC heterozygotes and CC homozygotes, indicating a functional role of the associated CD95 variant, possibly in supporting lymphocyte apoptosis.     

Plus- and Minus-End Directed Microtubule Motors Bind Simultaneously to Herpes Simplex Virus Capsids Using Different Inner Tegument Structures

Many viruses depend on host microtubule motors to reach their destined intracellular location. Viral particles of neurotropic alphaherpesviruses such as herpes simplex virus 1 (HSV1) show bidirectional transport towards the cell center as well as the periphery, indicating that they utilize microtubule motors of opposing directionality. To understand the mechanisms of specific motor recruitment, it is necessary to characterize the molecular composition of such motile viral structures. We have generated HSV1 capsids with different surface features without impairing their overall architecture, and show that in a mammalian cell-free system the microtubule motors dynein and kinesin-1 and the dynein cofactor dynactin could interact directly with capsids independent of other host factors. The capsid composition and surface was analyzed with respect to 23 structural proteins that are potentially exposed to the cytosol during virus assembly or cell entry. Many of these proteins belong to the tegument, the hallmark of all herpesviruses located between the capsid and the viral envelope. Using immunoblots, quantitative mass spectrometry and quantitative immunoelectron microscopy, we show that capsids exposing inner tegument proteins such as pUS3, pUL36, pUL37, ICP0, pUL14, pUL16, and pUL21 recruited dynein, dynactin, kinesin-1 and kinesin-2. In contrast, neither untegumented capsids exposing VP5, VP26, pUL17 and pUL25 nor capsids covered by outer tegument proteins such as vhs, pUL11, ICP4, ICP34.5, VP11/12, VP13/14, VP16, VP22 or pUS11 bound microtubule motors. Our data suggest that HSV1 uses different structural features of the inner tegument to recruit dynein or kinesin-1. Individual capsids simultaneously accommodated motors of opposing directionality as well as several copies of the same motor. Thus, these associated motors either engage in a tug-of-war or their activities are coordinately regulated to achieve net transport either to the nucleus during cell entry or to cytoplasmic membranes for envelopment during assembly.     

Characterizing the socially transmitted foraging tactic “sponging” by bottlenose dolphins (Tursiops sp.) in the western gulf of Shark Bay,Western Australia

Individual foraging tactics are widespread in animals and have ecological and evolutionary implications. Indo‐Pacific bottlenose dolphins (Tursiops sp.) in Shark Bay, Western Australia, exhibit a foraging tactic involving tool use, called “sponging.” Sponging is vertically, socially transmitted through the matriline and, to date, has been described in detail in the eastern gulf of Shark Bay (ESB). Here, we characterize sponging in the western gulf of Shark Bay (WSB), in which a different matriline engages in the behavior. We identified 40 individual “spongers” in 9 mo of boat‐based surveys over three field seasons. As is the case in ESB, the majority of WSB spongers was female and engaged in sponging in deep channel habitats. In contrast to ESB, however, there was no difference in the number of associates between spongers and nonspongers in WSB, and activity budgets differed between spongers and deep‐water nonspongers; spongers foraged more frequently and rested less than nonspongers. Group sizes in deep channel habitat, where sponging was prevalent, were typically larger than those in shallow habitat, except for foraging, perhaps indicative of higher predator abundance and/or scattered prey distribution in deep‐water habitat. This research improves our understanding of within‐population foraging variations in bottlenose dolphins.     

Effects of genetic variants in ADCY5, GIPR, GCKR and VPS13C on early impairment of glucose and insulin metabolism in children

Objective

Recent genome-wide association studies identified novel candidate genes for fasting and 2 h blood glucose and insulin levels in adults. We investigated the role of four of these loci (ADCY5, GIPR, GCKR and VPS13C) in early impairment of glucose and insulin metabolism in children.

Research Design and Methods

We genotyped four variants (rs2877716; rs1260326; rs10423928; rs17271305) in 638 Caucasian children with detailed metabolic testing including an oGTT and assessed associations with measures of glucose and insulin metabolism (including fasting blood glucose, insulin levels and insulin sensitivity/secretion indices) by linear regression analyses adjusted for age, sex, BMI-SDS and pubertal stage.

Results

The major allele (C) of rs2877716 (ADCY5) was nominally associated with decreased fasting plasma insulin (P = 0.008), peak insulin (P = 0.009) and increased QUICKI (P = 0.016) and Matsuda insulin sensitivity index (P = 0.013). rs17271305 (VPS13C) was nominally associated with 2 h blood glucose (P = 0.009), but not with any of the insulin or insulin sensitivity parameters. We found no association of the GIPR and GCKR variants with parameters of glucose and insulin metabolism. None of the variants correlated with anthropometric traits such as height, WHR or BMI-SDS, which excluded potential underlying associations with obesity.

Conclusions

Our data on obese children indicate effects of genetic variation within ADCY5 in early impairment of insulin metabolism and VPS13C in early impairment of blood glucose homeostasis.     

Langfristige Auswirkungen ehemaliger Bewirtschaftungsvarianten auf die Diversit?t von Blütenpflanzen, Moosen und Flechten eines Kalkmagerrasens

Jeschke M., Kiehl K., Pfadenhauer J. and Gigon A. 2008. Long-term effects of former management on the diversity of vascular plants, mosses and lichens in a calcareous grassland. Bot. Helv. 118: 95 – 109. Long-term effects of grassland management on the species richness of vascular plants, mosses and lichens were studied in a calcareous grassland near Schaffhausen (Switzerland). Experimental plots were treated for 22 years (1977 – 1999) with different mowing regimes, annual burning in late winter, or abandonment. From 2000 onwards, all plots were mown annually in autumn. Five years later (2004), we studied the after-effects of the previous treatments by recording the cover of vascular plants, mosses and lichens in nested plots with sizes from 0.01 m² to 16 m². Vascular plant species richness as well as the number of species characteristic for calcareous grasslands were significantly lower in formerly abandoned or burnt plots than in regularly mown plots, with larger differences on small subplots. There was no significant effect of the previous mowing time (July vs. October) or mowing frequency (annual or every two years) on vascular plants. Mosses and lichens had been eliminated by annual burning between 1977 and 1999, and only few moss species re-colonized the formerly burnt plots until 2004. The species richness of mosses did not differ between formerly abandoned and mown plots, but there were more mesophytic and fewer xerophytic species in the formerly abandoned plots. Mesophytic mosses were also more abundant in plots that used to be mown every second year than in those mown annually. Our data indicate that even after five years of similar management, i.e. annual mowing in autumn, the previous long-term management had strong after-effects on species richness and species composition of calcareous grasslands. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Eingereicht am 19. M?rz 2008, Angenommen am 30. August 2008, Redaktion: Sabine Güsewell     

Equivalence of regression curves sharing common parameters

In clinical trials, the comparison of two different populations is a common problem. Nonlinear (parametric) regression models are commonly used to describe the relationship between covariates, such as concentration or dose, and a response variable in the two groups. In some situations, it is reasonable to assume some model parameters to be the same, for instance, the placebo effect or the maximum treatment effect. In this paper, we develop a (parametric) bootstrap test to establish the similarity of two regression curves sharing some common parameters. We show by theoretical arguments and by means of a simulation study that the new test controls its significance level and achieves a reasonable power. Moreover, it is demonstrated that under the assumption of common parameters, a considerably more powerful test can be constructed compared with the test that does not use this assumption. Finally, we illustrate the potential applications of the new methodology by a clinical trial example.     

Gamma-aminobutyric acid type B receptors are constitutively internalized via the clathrin-dependent pathway and targeted to lysosomes for degradation

Receptor internalization is recognized as an important mechanism for rapidly regulating cell surface numbers of receptors. However, there are conflicting results on the existence of rapid endocytosis of gamma-aminobutyric acid, type B (GABAB) receptors. Therefore, we analyzed internalization of GABAB receptors expressed in HEK 293 cells qualitatively and quantitatively using immunocytochemical, cell surface enzyme-linked immunosorbent assay, and biotinylation methods. The data indicate the existence of rapid constitutive receptor internalization, with the first endocytosed receptors being observed in proximity of the plasma membrane after 10 min. After 120 min, a loss of about 40-50% of cell surface receptors was detected. Stimulation of GABAB receptors with GABA or baclofen did not enhance endocytosis of receptors, indicating the lack of agonist-induced internalization. The data suggest that GABAB receptors were endocytosed via the classical dynamin- and clathrin-dependent pathway and accumulated in an endosomal sorting compartment before being targeted to lysosomes for degradation. No evidence for recycling of receptors back to the cell surface was found. In conclusion, the results indicate the presence of constitutive internalization of GABAB receptors via clathrin-coated pits, which resulted in lysosomal degradation of the receptors.     

A family of modular type mannuronan C-5-epimerase genes controls alginate structure in Azotobacter vinelandii

The ToxR protein is a transmembrane protein that regulates the expression of several virulence factors of Vibrio cholerae. Previous analysis of fusion proteins between ToxR and alkaline phosphatase (ToxR-PhoA) suggested that ToxR was active as a dimer. In order to determine whether dimerization of the ToxR periplasmic domain was essential for activity, this domain was replaced by monomeric and dimeric protein domains. Surprisingly, PhoA (dimeric), β-lactamase (monomeric, ToxR–Bla), or the leucine zipper of GCN4 (dimeric, ToxR-GCN4-M) could substitute functionally for the ToxR periplasmic domain. ToxR-GCN4 fusion proteins, in which the ToxR trans-membrane domain was eliminated (ToxR-GCN4-C), were inactive, but an additional fusion protein that contained a heterologous membrane-spanning domain retained activity. Strains containing each of these ToxR fusion proteins were analysed for in vivo colonization properties and response to in vitro growth conditions that are known to affect expression of the ToxR regulon. Strains containing ToxR-GCN4-M and ToxR-Bla responded like wild-type strains to in vitro growth conditions. In the infant-mouse colonization model, strains containing ToxR fusion proteins were all deficient in colonization relative to strains containing wild-type ToxR, and strains containing monomeric ToxR-Bla were most severely outcompeted. These results suggest that, under in vitro conditions, ToxR does not require a dimerized periplasmic domain, but that, under in vivo conditions, the correct conformation of the ToxR periplasmic domain may be more important for function.