A systems biology approach to investigate the response of Synechocystis sp. PCC6803 to a high salt environment.

BACKGROUND: Salt overloading during agricultural processes is causing a decrease in crop productivity due to saline sensitivity. Salt tolerant cyanobacteria share many cellular characteristics with higher plants and therefore make ideal model systems for studying salinity stress. Here, the response of fully adapted Synechocystis sp. PCC6803 cells to the addition of 6% w/v NaCl was investigated using proteomics combined with targeted analysis of transcripts. RESULTS: Isobaric mass tagging of peptides led to accurate relative quantitation and identification of 378 proteins, and approximately 40% of these were differentially expressed after incubation in BG-11 media supplemented with 6% salt for 9 days. Protein abundance changes were related to essential cellular functional alterations. Differentially expressed proteins involved in metabolic responses were also analysed using the probabilitistic tool Mixed Model on Graphs (MMG), where the role of energy conversion through glycolysis and reducing power through pentose phosphate pathway were highlighted. Temporal RT-qPCR experiments were also run to investigate protein expression changes at the transcript level, for 14 non-metabolic proteins. In 9 out of 14 cases the mRNA changes were in accordance with the proteins. CONCLUSION: Synechocystis sp. PCC6803 has the ability to regulate essential metabolic processes to enable survival in high salt environments. This adaptation strategy is assisted by further regulation of proteins involved in non-metabolic cellular processes, supported by transcriptional and post-transcriptional control. This study demonstrates the effectiveness of using a systems biology approach in answering environmental, and in particular, salt adaptation questions in Synechocystis sp. PCC6803

Detection and monitoring of high stress concentration zones induced by coal mining using numerical and microseismic method

Zones of high stress concentration induced by coal mining at a depth of 1250 meters in the Lorraine Collieries are detected and monitored using a combination of numerical and microseismic methods. Changes in the stress state induced by coal mining are estimated by means of numerical simulations. The areas of high stress concentration are located and monitored by local microseismic network. The study of microseismic activity recorded during mining made it possible to localise the zones of high stress concentration and validate the calculations done by numerical modelling. The results of this study hold out interesting prospects for using a combination of numerical modelling and microseismic monitoring for detecting, locating and monitoring of the zones prone to rockbursts

Reduced keratin expression in colorectal neoplasia and associated fields is reversible by diet and resection

Abstract Background Patients with adenomatous colonic polyps are at increased risk of developing further polyps suggesting field-wide alterations in cancer predisposition. The current study aimed to identify molecular alterations in the normal mucosa in the proximity of adenomatous polyps and to assess the modulating effect of butyrate, a chemopreventive compound produced by fermentation of dietary residues. Methods A cross-sectional study was undertaken in patients with adenomatous polyps: biopsy samples were taken from the adenoma, and from macroscopically normal mucosa on the contralateral wall to the adenoma and from the mid-sigmoid colon. In normal subjects biopsies were taken from the mid-sigmoid colon. Biopsies were frozen for proteomic analysis or formalin-fixed for immunohistochemistry. Proteomic analysis was undertaken using iTRAQ workflows followed by bioinformatics analyses. A second dietary fibre intervention study arm used the same endpoints and sampling strategy at the beginning and end of a high-fibre intervention. Results Key findings were that keratins 8, 18 and 19 were reduced in expression level with progressive proximity to the lesion. Lesional tissue exhibited multiple K8 immunoreactive bands and overall reduced levels of keratin. Biopsies from normal subjects with low faecal butyrate also showed depressed keratin expression. Resection of the lesion and elevation of dietary fibre intake both appeared to restore keratin expression level. Conclusion Changes in keratin expression associate with progression towards neoplasia, but remain modifiable risk factors. Dietary strategies may improve secondary chemoprevention. Trial registration number ISRCTN90852168. Keywords: ADENOMA, BUTYRATE, CYTOKERATINS, DIETARY FIBR

The quantitative proteomic response of Synechocystis sp. PCC6803 to phosphate acclimation.

BACKGROUND: Inorganic phosphate (Pi) is a critical nutrient for all life and is periodically limiting in marine and freshwater provinces, yet little is understood how organisms acclimate to fluctuations in Pi within their environment. To investigate whole cell adaptation, we grew Synechocystis sp. PCC6803, a model freshwater cyanobacterium, in 3%, and 0.3% inorganic phosphate (Pi) media. The cells were allowed to acclimate over 60 days, and cells were harvested for quantitative high throughput mass spectrometry-based proteomics using the iTRAQ™ labelling technology. RESULTS: In total, 120 proteins were identified, and 52 proteins were considered differentially abundant compared to the control. Alkaline phosphatase (APase) activities correlated significantly (p < 0.05) with observed relative PhoA abundances. PstS1 and PstS2 were both observed, yet PstS1 was not differentially more abundant than the control. Phycobilisome protein abundances appeared to be coordinated, and are significantly less abundant in 0.3% Pi than 3% Pi cultures. Also, the central metabolic cell function appears to have shifted towards the production of (NADPH) reducing energy and nucleotide sugars. CONCLUSIONS: This acclimation response bears strong similarity to the previously reported response to nitrogen deprivation within Synechocystis sp. PCC 6803. However, it also demonstrates some characteristics of desiccation stress, such as the regulation of fatty acids and increased abundance of rehydrin in the 3% Pi culture

The HLA-B*57:01 allele corresponds to a very large MHC haploblock likely explaining its massive effect for HIV-1 elite control

IntroductionWe have reanalyzed the genomic data of the International Collaboration for the Genomics of HIV (ICGH), centering on HIV-1 Elite Controllers.MethodsWe performed a genome-wide Association Study comparing 543 HIV Elite Controllers with 3,272 uninfected controls of European descent. Using the latest database for imputation, we analyzed 35,552 Single Nucleotide Polymorphisms (SNPs) within the Major Histocompatibility Complex (MHC) region.ResultsOur analysis identified 2,626 SNPs significantly associated (p&lt;5. 10-8) with elite control of HIV-1 infection, including well-established MHC signals such as the rs2395029-G allele which tags HLA-B*57:01. A thorough investigation of SNPs in linkage disequilibrium with rs2395029 revealed an extensive haploblock spanning 1.9 megabases in the MHC region tagging HLA-B*57:01, comprising 379 SNP alleles impacting 72 genes. This haploblock contains damaging variations in proteins like NOTCH4 and DXO and is also associated with a strong differential pattern of expression of multiple MHC genes such as HLA-B, MICB, and ZBTB12. The study was expanded to include two cohorts of seropositive African-American individuals, where a haploblock tagging the HLA-B*57:03 allele was similarly associated with control of viral load. The mRNA expression profile of this haploblock in African Americans closely mirrored that in the European cohort.DiscussionThese findings suggest that additional molecular mechanisms beyond the conventional antigen-presenting role of class I HLA molecules may contribute to the observed influence of HLA-B*57:01/B*57:03 alleles on HIV-1 elite control. Overall, this study has uncovered a large haploblock associated with HLA-B*57 alleles, providing novel insights into their massive effect on HIV-1 elite control

Quantitative proteomic analysis of the influence of lignin on biofuel production by Clostridium acetobutylicum ATCC 824

Background: Clostridium acetobutylicum has been a focus of research because of its ability to produce high-value compounds that can be used as biofuels. Lignocellulose is a promising feedstock, but the lignin–cellulose–hemicellulose biomass complex requires chemical pre-treatment to yield fermentable saccharides, including cellulose-derived cellobiose, prior to bioproduction of acetone–butanol–ethanol (ABE) and hydrogen. Fermentation capability is limited by lignin and thus process optimization requires knowledge of lignin inhibition. The effects of lignin on cellular metabolism were evaluated for C. acetobutylicum grown on medium containing either cellobiose only or cellobiose plus lignin. Microscopy, gas chromatography and 8-plex iTRAQ-based quantitative proteomic technologies were applied to interrogate the effect of lignin on cellular morphology, fermentation and the proteome. Results: Our results demonstrate that C. acetobutylicum has reduced performance for solvent production when lignin is present in the medium. Medium supplemented with 1 g L−1 of lignin led to delay and decreased solvents production (ethanol; 0.47 g L−1 for cellobiose and 0.27 g L−1 for cellobiose plus lignin and butanol; 0.13 g L−1 for cellobiose and 0.04 g L−1 for cellobiose plus lignin) at 20 and 48 h, respectively, resulting in the accumulation of acetic acid and butyric acid. Of 583 identified proteins (FDR < 1 %), 328 proteins were quantified with at least two unique peptides. Up- or down-regulation of protein expression was determined by comparison of exponential and stationary phases of cellobiose in the presence and absence of lignin. Of relevance, glycolysis and fermentative pathways were mostly down-regulated, during exponential and stationary growth phases in presence of lignin. Moreover, proteins involved in DNA repair, transcription/translation and GTP/ATP-dependent activities were also significantly affected and these changes were associated with altered cell morphology. Conclusions: This is the first comprehensive analysis of the cellular responses of C. acetobutylicum to lignin at metabolic and physiological levels. These data will enable targeted metabolic engineering strategies to optimize biofuel production from biomass by overcoming limitations imposed by the presence of lignin

A Global Overview of the Genetic and Functional Diversity in the Helicobacter pylori cag Pathogenicity Island

The Helicobacter pylori cag pathogenicity island (cagPAI) encodes a type IV secretion system. Humans infected with cagPAI–carrying H. pylori are at increased risk for sequelae such as gastric cancer. Housekeeping genes in H. pylori show considerable genetic diversity; but the diversity of virulence factors such as the cagPAI, which transports the bacterial oncogene CagA into host cells, has not been systematically investigated. Here we compared the complete cagPAI sequences for 38 representative isolates from all known H. pylori biogeographic populations. Their gene content and gene order were highly conserved. The phylogeny of most cagPAI genes was similar to that of housekeeping genes, indicating that the cagPAI was probably acquired only once by H. pylori, and its genetic diversity reflects the isolation by distance that has shaped this bacterial species since modern humans migrated out of Africa. Most isolates induced IL-8 release in gastric epithelial cells, indicating that the function of the Cag secretion system has been conserved despite some genetic rearrangements. More than one third of cagPAI genes, in particular those encoding cell-surface exposed proteins, showed signatures of diversifying (Darwinian) selection at more than 5% of codons. Several unknown gene products predicted to be under Darwinian selection are also likely to be secreted proteins (e.g. HP0522, HP0535). One of these, HP0535, is predicted to code for either a new secreted candidate effector protein or a protein which interacts with CagA because it contains two genetic lineages, similar to cagA. Our study provides a resource that can guide future research on the biological roles and host interactions of cagPAI proteins, including several whose function is still unknown

iTRAQ Identification of Candidate Serum Biomarkers Associated with Metastatic Progression of Human Prostate Cancer

A major challenge in the management of patients with prostate cancer is identifying those individuals at risk of developing metastatic disease, as in most cases the disease will remain indolent. We analyzed pooled serum samples from 4 groups of patients (n = 5 samples/group), collected prospectively and actively monitored for a minimum of 5 yrs. Patients groups were (i) histological diagnosis of benign prostatic hyperplasia with no evidence of cancer ‘BPH’, (ii) localised cancer with no evidence of progression, ‘non-progressing’ (iii) localised cancer with evidence of biochemical progression, ‘progressing’, and (iv) bone metastasis at presentation ‘metastatic’. Pooled samples were immuno-depleted of the 14 most highly abundant proteins and analysed using a 4-plex iTRAQ approach. Overall 122 proteins were identified and relatively quantified. Comparisons of progressing versus non-progressing groups identified the significant differential expression of 25 proteins (p<0.001). Comparisons of metastatic versus progressing groups identified the significant differential expression of 23 proteins. Mapping the differentially expressed proteins onto the prostate cancer progression pathway revealed the dysregulated expression of individual proteins, pairs of proteins and ‘panels’ of proteins to be associated with particular stages of disease development and progression. The median immunostaining intensity of eukaryotic translation elongation factor 1 alpha 1 (eEF1A1), one of the candidates identified, was significantly higher in osteoblasts in close proximity to metastatic tumour cells compared with osteoblasts in control bone (p = 0.0353, Mann Whitney U). Our proteomic approach has identified leads for potentially useful serum biomarkers associated with the metastatic progression of prostate cancer. The panels identified, including eEF1A1 warrant further investigation and validation

CyanoFactory, a European consortium to develop technologies needed to advance cyanobacteria as chassis for production of chemicals and fuels

CyanoFactory, Design, construction and demonstration of solar biofuel production using novel (photo)synthetic cell factories, was an R&D project developed in response to the European Commission FP7-ENERGY-2012-1 call “Future Emerging Technologies” and the need for significant advances in both new science and technologies to convert solar energy into a fuel. CyanoFactory was an example of “purpose driven” research and development with identified scientific goals and creation of new technologies. The present overview highlights significant outcomes of the project, three years after its successful completion. The scientific progress of CyanoFactory involved: (i) development of a ToolBox for cyanobacterial synthetic biology; (ii) construction of DataWarehouse/Bioinformatics web-based capacities and functions; (iii) improvement of chassis growth, functionality and robustness; (iv) introduction of custom designed genetic constructs into cyanobacteria, (v) improvement of photosynthetic efficiency towards hydrogen production; (vi) biosafety mechanisms; (vii) analyses of the designed cyanobacterial cells to identify bottlenecks with suggestions on further improvements; (viii) metabolic modelling of engineered cells; (ix) development of an efficient laboratory scale photobioreactor unit; and (x) the assembly and experimental performance assessment of a larger (1350 L) outdoor flat panel photobioreactor system during two seasons. CyanoFactory - Custom design and purpose construction of microbial cells for the production of desired products using synthetic biology – aimed to go beyond conventional paths to pursue innovative and high impact goals. CyanoFactory brought together ten leading European partners (universities, research organizations and enterprises) with a common goal – to develop the future technologies in Synthetic biology and Advanced photobioreactors