Role of phosphatidylglycerol in the function and assembly of Photosystem II reaction center, studied in a cdsA-inactivated PAL mutant strain of Synechocystis sp. PCC6803 that lacks phycobilisomes

To analyze the role of phosphatidylglycerol (PG) in photosynthetic membranes of cyanobacteria we used two mutants of Synechocystis sp. PCC6803: the PAL mutant which has no phycobilisomes and shows a high PSII/PSI ratio, and a mutant derived from it by inactivating its cdsA gene encoding cytidine 5'-diphosphate diacylglycerol synthase, a key enzyme in PG synthesis. In a medium supplemented with PG the PAL/DeltacdsA mutant cells grew photoautotrophically. Depletion of PG in the medium resulted (a) in an arrest of cell growth and division, (b) in a slowdown of electron transfer from the acceptor Q(A) to Q(B) in PSII and (c) in a modification of chlorophyll fluorescence curve. The depletion of PG affected neither the redox levels of Q(A) nor the S(2) state of the oxygen-evolving manganese complex, as indicated by thermoluminescence studies. Two-dimensional PAGE showed that in the absence of PG (a) the PSII dimer was decomposed into monomers, and (b) the CP43 protein was detached from a major part of the PSII core complex. [(35)S]-methionine labeling confirmed that PG depletion did not block de novo synthesis of the PSII proteins. We conclude that PG is required for the binding of CP43 within the PSII core complex.     

The mapping of linear B‐cell epitope regions in the extracellular parts of the desmoglein 1 and 3 proteins: recognition of immobilized peptides by pemphigus patients' serum autoantibodies

Desmosomal transmembrane glycoproteins desmogleins (Dsg) 1 and 3 are targets of life‐threatening autoimmune blistering disorders such as Pemphigus vulgaris (PV) and Pemphigus foliaceus (PF). In these diseases, pemphigus autoantibodies are produced against Dsg1 and Dsg3 proteins. The autoantibodies bind to these transmembrane elements leading to a loss of desmosomal cell–cell adhesion and clinically, to the presence of blisters and erosions. Identification, characterization, and detailed analysis of the binding sites of autoantibodies have an outstanding importance in understanding the immunopathology of the disease and also in the design of novel diagnostics. Here, we describe the localization of the B‐cell epitope regions of Dsg1 and Dsg3 proteins' extracellular parts recognized by IgG‐type serum autoantibodies of patients with PV and PF. In our study, overlapping pentadecapeptides were synthesized on hydroxypropyl methacrylate pins based on the results of in silico predictions. To detect the interaction between the serum autoantibodies and the immobilized synthetic peptides, modified Enzyme Linked Immunosorbent Assay (ELISA) was performed with pin‐attached peptides testing the serum samples of ten patients and four healthy donors. We identified five possible epitope regions (aa86‐110, aa196‐220, aa226‐250, aa326‐340, and aa486‐520) within the extracellular part of the Dsg1 and four possible epitope regions (aa64‐78, aa330‐344, aa375‐399, and aa446‐460) within that of the Dsg3 protein sequence using these methods. Our data showed that serum autoantibodies of patients, previously identified as Dsg1 and Dsg3 positive, are able to recognize continuous linear epitope regions of both Dsg1 and Dsg3 proteins using pin‐bound overlapping peptides in modified ELISAs. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Unique picoeukaryotic algal community under multiple environmental stress conditions in a shallow, alkaline pan

Winter phytoplankton communities in the shallow alkaline pans of Hungary are frequently dominated by picoeukaryotes, sometimes in particularly high abundance. In winter 2012, the ice-covered alkaline Zab-szék pan was found to be extraordinarily rich in picoeukaryotic green algae (42–82 × 10⁶ cells ml^?1) despite the simultaneous presence of multiple stressors (low temperature and light intensity with high pH and salinity). The maximum photosynthetic rate of the picoeukaryote community was 1.4 μg C μg chlorophyll a ^?1 h^?1 at 125 μmol m^?2 s^?1. The assimilation rates compared with the available light intensity measured on the field show that the community was considerably light-limited. Estimated areal primary production was 180 mg C m^?2 d^?1. On the basis of the 18S rRNA gene analysis (cloning and DGGE), the community was phylogenetically heterogeneous with several previously undescribed chlorophyte lineages, which indicates the ability of picoeukaryotic communities to maintain high genetic diversity under extreme conditions.     

Right Ventricular Adaptation Is Associated with the Glu298Asp Variant of the NOS3 Gene in Elite Athletes

Nitric oxide (NO), an important endogenous pulmonary vasodilator is synthetized by the endothelial NO synthase (NOS3). Reduced NO bioavailability and thus the Glu298Asp polymorphism of NOS3 may enhance right ventricular (RV) afterload and hypertrophic remodeling and influence athletic performance. To test this hypothesis world class level athletes (water polo players, kayakers, canoeists, rowers, swimmers, n = 126) with a VO₂ maximum greater than 50ml/kg/min were compared with non-athletic volunteers (n = 155). Cardiopulmonary exercise tests and cardiac magnetic resonance imaging (cMRI) were performed to determine structural or functional changes. Genotype distribution of the NOS3 Glu298Asp polymorphism was not affected by gender or physical performance. Cardiac MRI showed increased stroke volume with eccentric hypertrophy in all athletes regardless of their genotype. However, the Asp allelic variant carriers had increased RV mass index (32±6g versus 27±6g, p<0.01) and larger RV stroke volume index (71±10ml versus 64±10ml, p<0.01) than athletes with a Glu/Glu genotype. Genotype was not significantly associated with athletic performance. In the non-athletic group no genotype related differences were detected. The association between the NOS3 Glu298Asp polymorphism and RV structure and dimension in elite athletes emphasizes the importance of NOS3 gene function and NO bioavailability in sport related cardiac adaptation.     

Targeting cellular metabolism using rapamycin and/or doxycycline enhances anti-tumour effects in human glioma cells

Background

Glioma is the most common highly aggressive, primary adult brain tumour. Clinical data show that therapeutic approaches cannot reach the expectations in patients, thus gliomas are mainly incurable diseases. Tumour cells can adapt rapidly to alterations during therapeutic treatments related to their metabolic rewiring and profound heterogeneity in tissue environment. Renewed interests aim to develop effective treatments targeting angiogenesis, kinase activity and/or cellular metabolism. mTOR (mammalian target of rapamycin), whose hyper-activation is characteristic for many tumours, promotes metabolic alterations, macromolecule biosynthesis, cellular growth and survival. Unfortunately, mTOR inhibitors with their lower toxicity have not resulted in appreciable survival benefit. Analysing mTOR inhibitor sensitivity, other metabolism targeting treatments and their combinations could help to find potential agents and biomarkers for therapeutic development in glioma patients.

Methods

In vitro proliferation assays, protein expression and metabolite concentration analyses were used to study the effects of mTOR inhibitors, other metabolic treatments and their combinations in glioma cell lines. Furthermore, mTOR activity and cellular metabolism related protein expression patterns were also investigated by immunohistochemistry in human biopsies. Temozolomide and/or rapamycin treatments altered the expressions of enzymes related to lipid synthesis, glycolysis and mitochondrial functions as consequences of metabolic adaptation; therefore, other anti-metabolic drugs (chloroquine, etomoxir, doxycycline) were combined in vitro.

Results

Our results suggest that co-targeting metabolic pathways had tumour cell dependent additive/synergistic effects related to mTOR and metabolic protein expression patterns cell line dependently. Drug combinations, especially rapamycin?+?doxycycline may have promising anti-tumour effect in gliomas. Additionally, our immunohistochemistry results suggest that metabolic and mTOR activity alterations are not related to the recent glioma classification, and these protein expression profiles show individual differences in patients’ materials.

Conclusions

Based on these, combinations of different new/old drugs targeting cellular metabolism could be promising to inhibit high adaptation capacity of tumour cells depending on their metabolic shifts. Relating to this, such a development of current therapy needs to find special biomarkers to characterise metabolic heterogeneity of gliomas.

     

Late Quaternary woolly mammoth (Mammuthus primigenius Blum) remains from southern Transdanubia,Hungary

Six samples of subfossil tusk, bone and tooth remains from the woolly mammoth (Mammuthus primigenius Blum) were discovered in south-western Hungary. The remains are relatively well preserved in a Late Pleistocene loess deposit. The samples have been radiocarbon dated (AMS) and are of Late Weichselian (MIS 2) age (21.8–24.1 ka cal BP). The skull fragments, the tusks and maxillary teeth are in close proximity to associated postcranial remains, indicating that the mammoth died where it was found. The size and characteristics of skeletal elements have allowed us to determine that this was a mature male of about 38 years of age.     

Genomic signatures of strain selection and enhancement in Bacillus atrophaeus var. globigii, a historical biowarfare simulant

Background

Despite the decades-long use of Bacillus atrophaeus var. globigii (BG) as a simulant for biological warfare (BW) agents, knowledge of its genome composition is limited. Furthermore, the ability to differentiate signatures of deliberate adaptation and selection from natural variation is lacking for most bacterial agents. We characterized a lineage of BGwith a long history of use as a simulant for BW operations, focusing on classical bacteriological markers, metabolic profiling and whole-genome shotgun sequencing (WGS).

Results

Archival strains and two “present day” type strains were compared to simulant strains on different laboratory media. Several of the samples produced multiple colony morphotypes that differed from that of an archival isolate. To trace the microevolutionary history of these isolates, we obtained WGS data for several archival and present-day strains and morphotypes. Bacillus-wide phylogenetic analysis identified B. subtilis as the nearest neighbor to B. atrophaeus. The genome of B. atrophaeus is, on average, 86% identical to B. subtilis on the nucleotide level. WGS of variants revealed that several strains were mixed but highly related populations and uncovered a progressive accumulation of mutations among the “military” isolates. Metabolic profiling and microscopic examination of bacterial cultures revealed enhanced growth of “military” isolates on lactate-containing media, and showed that the “military” strains exhibited a hypersporulating phenotype.

Conclusions

Our analysis revealed the genomic and phenotypic signatures of strain adaptation and deliberate selection for traits that were desirable in a simulant organism. Together, these results demonstrate the power of whole-genome and modern systems-level approaches to characterize microbial lineages to develop and validate forensic markers for strain discrimination and reveal signatures of deliberate adaptation.     

Mite species inhabiting commercial bumblebee (<Emphasis Type="Italic">Bombus terrestris</Emphasis>) nests in Polish greenhouses

Nests of social insects are usually inhabited by various mite species that feed on pollen, other micro-arthropods or are parasitic. Well-known negative effects of worldwide economic importance are caused by mites parasitizing honeybee colonies. Lately, attention has focused on the endoparasitic mite Locustacarus buchneri that has been found in commercial bumblebees. However, little is known of other mites associated with commercial bumblebee nests. Transportation of commercial bumblebee colonies with unwanted residents may introduce foreign mite species to new localities. In this study, we assessed the prevalence and species composition of mites associated with commercial bumblebee nests and determined if the mites are foreign species for Poland and for Europe. The study was conducted on 37 commercial bumblebee nests from two companies (Dutch and Israeli), originating from two greenhouses in southern Poland, and on 20 commercial bumblebee colonies obtained directly from suppliers. The species composition and abundance of mites inhabiting commercial bumblebee nests were determined. Seven mite species from three families were found in nests after greenhouse exploitation. The predominant mite species was Tyrophagus putrescentiae (Acaridae) that was a 100-fold more numerous than representatives of the family Laelapidae (Hypoaspis marginepilosa, H. hyatti, H. bombicolens). Representatives of Parasitidae (Parasitellus fucorum, P. crinitus, P. ignotus) were least numerous. All identified mite species are common throughout Europe, foreign species were not found. Mites were not detected in nests obtained directly from suppliers. We conclude that probably bumblebee nests are invaded by local mite species during greenhouse exploitation.