Arabidopsis E2FA stimulates proliferation and endocycle separately through RBR-bound and RBR-free complexes

Post-embryonic growth in plants depends on the continuous supply of undifferentiated cells within meristems. Proliferating cells maintain their competence for division by active repression of differentiation and the associated endocycle entry. We show by upregulation and downregulation of E2FA that it is required for maintaining proliferation, as well as for endocycle entry. While E2FB-RBR1 (retinoblastoma-related protein 1) complexes are reduced after sucrose addition or at elevated CYCD3;1 levels, E2FA maintains a stable complex with RBR1 in proliferating cells. Chromatin immunoprecipitation shows that RBR1 binds in the proximity of E2F promoter elements in CCS52A1 and CSS52A2 genes, central regulators for the switch from proliferation to endocycles. Overexpression of a truncated E2FA mutant (E2FA(ΔRB)) lacking the RBR1-binding domain interferes with RBR1 recruitment to promoters through E2FA, leading to decreased meristem size in roots, premature cell expansion and hyperactivated endocycle in leaves. E2F target genes, including CCS52A1 and CCS52A2, are upregulated in E2FA(ΔRB) and e2fa knockout lines. These data suggest that E2FA in complex with RBR1 forms a repressor complex in proliferating cells to inhibit premature differentiation and endocycle entry. Thus, E2FA regulates organ growth via two distinct, sequentially operating pathways.     

Patterns of Fungal and Bacterial Carbon Mineralization Across Northern European Peatlands

The fungal and bacterial activity was determined in 20 northern European peatlands ranging from ombrotrophic bogs to eutrophic fens with key differences in degree of humification, pH, dry bulk density, carbon (C) content and vegetation communities using the selective inhibition (SI) technique. These peatlands were partly disturbed and the respective water tables lowered below the surface layer. Basal respiration ranged from 24 to 128 µg CO₂-C g^?1 dry peat d^?1. Bacterial contributions to CO₂ production were high in most peatlands and showed the following pattern: eutrophic >> transitional ≥ mesotrophic >> ombrotrophic peatland types. The fungal-to-bacterial (F:B) ratios varied substantially within peatland type, and this was mainly attributed to differences in peat botanical compositions and chemistry. The computed mean Inhibitor Additivity Ratio (IAR) was quite close to 1 to suggest that the SI techniques can be used to partition eukaryotic and prokaryotic activity in wide range of peatlands. Overall, basal respiration, microbial biomass-C, fungal and bacterial activities varied across the studied peatland types, and such differences could have consequences for C- and nutrient-cycling as well as how bogs and fens will respond to environmental changes.     

Pertussis toxin blocks melatonin-induced pigment aggregation inXenopus dermal melanophores

Summary The molecular mechanism of action for the pineal hormone melatonin was explored by testing melatonin interaction with the components of the hormone-sensitive adenylate cyclase complex in aXenopus dermal melanophore bioassay. Forskolin was employed to stimulate melanosome dispersion. The ability of melatonin to reverse forskolin-stimulated pigment dispersion was assessed, as was the effect of pertussis toxin on the ability of melatonin to aggregate dispersed pigment.Forskolin elicited dispersal of melanosomes in a dose dependent manner (EC₅₀=12 nM) in meninges from stage 52–56 tadpoles ofXenopus laevis. Maximal pigment dispersion was obtained with 100 nM forskolin. Melatonin reversed this effect of forskolin (EC₅₀=1.5 nM), causing pigment aggregation. Pertussis toxin blocked the melatonin-induced aggregation (EC₅₀=358 ng/ml).Prior treatment of the melanophore containing meningeal explants with pertussis toxin results in blockade of melatonin induced pigment aggregation. A 41 kDa pertussis toxin substrate is found in explant homogenates treated with³²P-NAD and pertussis toxin. The availability of this substrate is reduced by prior treatment of intact explants with pertussis toxin and depletion of melatonin responsiveness corresponds to depletion of the 41 kDa substrate. Together, these data suggest that melatonin action upon amphibian dermal melanosomes is mediated by a system requiring a protein similar to the regulatory protein Ni used by mammalian cells to mediate the action of hormones which inhibit adenylate cyclase through a cell surface receptor.Abbreviations MI melanophore index - MSH melanocyte stimulating hormone - FCS fetal calf serum     

I-MOVE multi-centre case control study 2010-11: overall and stratified estimates of influenza vaccine effectiveness in Europe

Background

In the third season of I-MOVE (Influenza Monitoring Vaccine Effectiveness in Europe), we undertook a multicentre case-control study based on sentinel practitioner surveillance networks in eight European Union (EU) member states to estimate 2010/11 influenza vaccine effectiveness (VE) against medically-attended influenza-like illness (ILI) laboratory-confirmed as influenza.

Methods

Using systematic sampling, practitioners swabbed ILI/ARI patients within seven days of symptom onset. We compared influenza-positive to influenza laboratory-negative patients among those meeting the EU ILI case definition. A valid vaccination corresponded to > 14 days between receiving a dose of vaccine and symptom onset. We used multiple imputation with chained equations to estimate missing values. Using logistic regression with study as fixed effect we calculated influenza VE adjusting for potential confounders. We estimated influenza VE overall, by influenza type, age group and among the target group for vaccination.

Results

We included 2019 cases and 2391 controls in the analysis. Adjusted VE was 52% (95% CI 30-67) overall (N = 4410), 55% (95% CI 29-72) against A(H1N1) and 50% (95% CI 14-71) against influenza B. Adjusted VE against all influenza subtypes was 66% (95% CI 15-86), 41% (95% CI -3-66) and 60% (95% CI 17-81) among those aged 0-14, 15-59 and ≥60 respectively. Among target groups for vaccination (N = 1004), VE was 56% (95% CI 34-71) overall, 59% (95% CI 32-75) against A(H1N1) and 63% (95% CI 31-81) against influenza B.

Conclusions

Results suggest moderate protection from 2010-11 trivalent influenza vaccines against medically-attended ILI laboratory-confirmed as influenza across Europe. Adjusted and stratified influenza VE estimates are possible with the large sample size of this multi-centre case-control. I-MOVE shows how a network can provide precise summary VE measures across Europe.     

Das Auftreten eines besonderen Typs von Golgivesikeln während der Sekundärwandbildung vonMicrasterias denticulata Bréb

Zusammenfassung Bei der DesmidiaceeMicrasterias denticulata vollzieht sich die Bildung der Primär- und Sekundärwand während bestimmter Phasen der Zellentwicklung. Primäre und sekundäre Wände können an Hand ihrer unterschiedlichen Texturen identifiziert werden. Um mögliche feinstrukturelle Faktoren zu finden, die bei der Bildung der Sekundärwand eine Rolle spielen könnten, wurden Zellen während der Sekundärwandbildung für elektronenmikroskopische Untersuchungen präpariert.Während der Sekundärwandbildung konnte ein sehr eigenartiger Typ diskusförmiger Vesikel im Cytoplasma beobachtet werden, der offenbar von den Dictyosomen produziert wird. Diese flachen Vesikel, die sackförmige Gebilde an ihren Rändern tragen, werden in der Phase der Sekundärwandbildung in die Plasmamembran inkorporiert. Die flachen Areale der Vesikel zeichnen sich durch eine besonders dicke Membran aus (160–200 Å), die auf der Vesikelinnenseite mit globulären Partikeln von ca. 200 Å Durchmesser besetzt ist. Nach der Fusion der Membran der flachen Vesikel mit der Plasmamembran gelangen diese Partikel an die Außenseite der Plasmamembran, die als Bildungsort für die Cellulosefibrillen der Sekundärwand angesehen wird. Die Bedeutung der flachen Vesikel und der globulären Partikel auf der Membranoberfläche werden in ihrem Verhältnis zur Bildung der Mikrofibrillen diskutiert.

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The Occurrence of a special type of golgi-vesicles during secondary wall formation inMicrasterias denticulata bréb

Summary In the desmidMicrasterias denticulata the formation of the primary and secondary cell wall occur during discrete phases of cell differentiation. The primary and secondary cell walls can be identified according to their particular textures. In order to determine fine structural factors possibly involved in the formation of the secondary cell wall, cells undergoing secondary wall formation were prepared for electron microscopic investigations. During secondary wall formation a very particular type of disc-like vesicles in the cytoplasm could be observed, appearently produced by the dictyosomes. These flat vesicles, carring sack-like structures at their edges, are found to be incorporated into the plasma membrane during the period of secondary wall formation. The flat areas of the vesicles are characterized by an unusually thick membrane (160–200 Å) that carries globular particles of about 200 Å diameter on the inner membrane surface. By fusion of the flat vesicle-membrane with the plasma membrane, the globular particles reach the outside of the plasma membrane. This is the site where the cellulose microfibrils of the secondary wall are synthesized. The significance of the flat vesicles and the globular particles on the membrane surface are discussed in relation to microfibril production.

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Für die großzügige Unterstützung unserer Arbeiten danken wir der Deutschen Forsungsgemeinschaft.     

Architecture of the active post‐translational Sec translocon

In eukaryotes, most secretory and membrane proteins are targeted by an N‐terminal signal sequence to the endoplasmic reticulum, where the trimeric Sec61 complex serves as protein‐conducting channel (PCC). In the post‐translational mode, fully synthesized proteins are recognized by a specialized channel additionally containing the Sec62, Sec63, Sec71, and Sec72 subunits. Recent structures of this Sec complex in the idle state revealed the overall architecture in a pre‐opened state. Here, we present a cryo‐EM structure of the yeast Sec complex bound to a substrate, and a crystal structure of the Sec62 cytosolic domain. The signal sequence is inserted into the lateral gate of Sec61α similar to previous structures, yet, with the gate adopting an even more open conformation. The signal sequence is flanked by two Sec62 transmembrane helices, the cytoplasmic N‐terminal domain of Sec62 is more rigidly positioned, and the plug domain is relocated. We crystallized the Sec62 domain and mapped its interaction with the C‐terminus of Sec63. Together, we obtained a near‐complete and integrated model of the active Sec complex.     

Heart Rate Variability Biofeedback Does Not Substitute for Asthma Steroid Controller Medication

Despite previous findings of therapeutic effects for heart rate variability biofeedback (HRVB) on asthma, it is not known whether HRVB can substitute either for controller or rescue medication, or whether it affects airway inflammation. Sixty-eight paid volunteer steroid naïve study participants with mild or moderate asthma were given 3 months of HRVB or a comparison condition consisting of EEG alpha biofeedback with relaxing music and relaxed paced breathing (EEG+), in a two-center trial. All participants received a month of intensive asthma education prior to randomization. Both treatment conditions produced similar significant improvements on the methacholine challenge test (MCT), asthma symptoms, and asthma quality of life (AQOL). MCT effects were of similar size to those of enhanced placebo procedures reported elsewhere, and were 65% of those of a course of a high-potency inhaled steroid budesonide given to a sub-group of participants following biofeedback training. Exhaled nitric oxide decreased significantly only in the HRVB group, 81% of the budesonide effect, but with no significant differences between groups. Participants reported becoming more relaxed during practice of both techniques. Administration of albuterol after biofeedback sessions produced a large improvement in pulmonary function test results, indicating that neither treatment normalized pulmonary function as a potent controller medication would have done. Impulse oscillometry showed increased upper airway (vocal cord) resistance during biofeedback periods in both groups. These data suggest that HRVB should not be considered an alternative to asthma controller medications (e.g., inhaled steroids), although both biofeedback conditions produced some beneficial effects, warranting further research, and suggesting potential complementary effects. Various hypotheses are presented to explain why HRVB effects on asthma appeared smaller in this study than in earlier studies. Clinical Trial Registration NCT02766374.     

Knowledge-based assessment of gene expression data from chemiluminescence detection.

The first problem in gene expression profiling to be solved is choosing the appropriate gene array, detection procedure, image analysis and data generation depending on the organism of interest, equipment and budget. The next one is how to deduce biologically meaningful data. We assessed gene expression data from chemiluminescent detection and empirically found criteria for the reliable identification of biologically meaningful expression ratios. Current statistical assessments are often applied unreflectedly concerning problems occurring in practice. So interesting results are considered to be irrelevant. This requires a laborious data check. We suggest automation. Our empirically found criteria were transformed into and validated by a knowledge-based system. This system is adaptable to all other methods of expression profiling. We compared the experience-based and new knowledge-based assessment of the expression data from our chemiluminescent and additionally radioactive detection of several experiments with published data to evaluate our entire procedure. With our adaptation of chemiluminescence detection to commercially available Escherichia coli gene arrays we present a useful alternative to common procedures in gene expression monitoring. Moreover, with our consideration of plasmid-harbouring E. coli strains we provide the opportunity to monitor gene expression during processes requiring any plasmids (e.g. recombinant protein expression).     

Integrative Omics Analysis of Rheumatoid Arthritis Identifies Non-Obvious Therapeutic Targets

Identifying novel therapeutic targets for the treatment of disease is challenging. To this end, we developed a genome-wide approach of candidate gene prioritization. We independently collocated sets of genes that were implicated in rheumatoid arthritis (RA) pathogenicity through three genome-wide assays: (i) genome-wide association studies (GWAS), (ii) differentially expression in RA fibroblast-like synoviocytes (FLS), and (iii) differentially methylation in RA FLS. Integrated analysis of these complementary data sets identified a significant enrichment of multi-evidence genes (MEGs) within pathways relating to RA pathogenicity. One MEG is Engulfment and Cell Motility Protein-1 (ELMO1), a gene not previously considered as a therapeutic target in RA FLS. We demonstrated in RA FLS that ELMO1 is: (i) expressed, (ii) promotes cell migration and invasion, and (iii) regulates Rac1 activity. Thus, we created links between ELMO1 and RA pathogenicity, which in turn validates ELMO1 as a potential RA therapeutic target. This study illustrated the power of MEG-based approaches for therapeutic target identification.