Whole Genome PCR Scanning Reveals the Syntenic Genome Structure of Toxigenic Vibrio cholerae Strains in the O1/O139 Population

Vibrio cholerae is commonly found in estuarine water systems. Toxigenic O1 and O139 V. cholerae strains have caused cholera epidemics and pandemics, whereas the nontoxigenic strains within these serogroups only occasionally lead to disease. To understand the differences in the genome and clonality between the toxigenic and nontoxigenic strains of V. cholerae serogroups O1 and O139, we employed a whole genome PCR scanning (WGPScanning) method, an rrn operon-mediated fragment rearrangement analysis and comparative genomic hybridization (CGH) to analyze the genome structure of different strains. WGPScanning in conjunction with CGH revealed that the genomic contents of the toxigenic strains were conservative, except for a few indels located mainly in mobile elements. Minor nucleotide variation in orthologous genes appeared to be the major difference between the toxigenic strains. rrn operon-mediated rearrangements were infrequent in El Tor toxigenic strains tested using I-CeuI digested pulsed-field gel electrophoresis (PFGE) analysis and PCR analysis based on flanking sequence of rrn operons. Using these methods, we found that the genomic structures of toxigenic El Tor and O139 strains were syntenic. The nontoxigenic strains exhibited more extensive sequence variations, but toxin coregulated pilus positive (TCP+) strains had a similar structure. TCP+ nontoxigenic strains could be subdivided into multiple lineages according to the TCP type, suggesting the existence of complex intermediates in the evolution of toxigenic strains. The data indicate that toxigenic O1 El Tor and O139 strains were derived from a single lineage of intermediates from complex clones in the environment. The nontoxigenic strains with non-El Tor type TCP may yet evolve into new epidemic clones after attaining toxigenic attributes

MCP-1 Upregulates Amylin Expression in Murine Pancreatic β Cells through ERK/JNK-AP1 and NF-κB Related Signaling Pathways Independent of CCR2

BACKGROUND: Amylin is the most abundant component of islet amyloid implicated in the development of type 2 diabetes. Plasma amylin levels are elevated in individuals with obesity and insulin resistance. Monocyte chemoattractant protein-1 (MCP-1, CCL2) is involved in insulin resistance of obesity and type 2 diabetes. We investigated the effect of MCP-1 on amylin expression and the underlying mechanisms with murine pancreatic β-cell line MIN6 and pancreatic islets. METHODOLOGY/PRINCIPAL FINDINGS: We found that MCP-1 induced amylin expression at transcriptional level and increased proamylin and intermediate forms of amylin at protein level in MIN6 cells and islets. However, MCP-1 had no effect on the expressions of proinsulin 1 and 2, as well as prohormone convertase (PC) 1/3 and PC2, suggesting that MCP-1 specifically induces amylin expression in β-cells. Mechanistic studies showed that although there is no detectable CCR2 mRNA in MIN6 cells and islets, pretreatment of MIN6 cells with pertussis toxin inhibited MCP-1 induced amylin expression, suggesting that alternative Gi-coupled receptor(s) mediates the inductive effect of MCP-1. MCP-1 rapidly induced ERK1/2 and JNK phosphorylation. Inhibitors for MEK1/2 (PD98059), JNK (SP600125) or AP1 (curcumin) significantly inhibited MCP-1-induced amylin mRNA expression. MCP-1 failed to induce amylin expression in pancreatic islets isolated from Fos knockout mice. EMSA showed that JNK and ERK1/2 were involved in MCP-1-induced AP1 activation. These results suggest that MCP-1 induces murine amylin expression through AP1 activation mediated by ERK1/2 or JNK. Further studies showed that treatment of MIN6 cells with NF-κB inhibitor or overexpression of IκBα dominant-negative construct in MIN6 cells significantly inhibited MCP-1-induced amylin expression, suggesting that NF-κB related signaling also participates in MCP-1-induced murine amylin expression. CONCLUSIONS/SIGNIFICANCE: MCP-1 induces amylin expression through ERK1/2/JNK-AP1 and NF-κB related signaling pathways independent of CCR2. Amylin upregulation by MCP-1 may contribute to elevation of plasma amylin in obesity and insulin resistance

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe

Novel genetic associations for blood pressure identified via gene-alcohol interaction in up to 570K individuals across multiple ancestries

Heavy alcohol consumption is an established risk factor for hypertension; the mechanism by which alcohol consumption impact blood pressure (BP) regulation remains unknown. We hypothesized that a genome-wide association study accounting for gene-alcohol consumption interaction for BP might identify additional BP loci and contribute to the understanding of alcohol-related BP regulation. We conducted a large two-stage investigation incorporating joint testing of main genetic effects and single nucleotide variant (SNV)-alcohol consumption interactions. In Stage 1, genome-wide discovery meta-analyses in approximate to 131 K individuals across several ancestry groups yielded 3,514 SNVs (245 loci) with suggestive evidence of association (P <1.0 x 10(-5)). In Stage 2, these SNVs were tested for independent external replication in individuals across multiple ancestries. We identified and replicated (at Bonferroni correction threshold) five novel BP loci (380 SNVs in 21 genes) and 49 previously reported BP loci (2,159 SNVs in 109 genes) in European ancestry, and in multi-ancestry meta-analyses (P < 5.0 x 10(-8)). For African ancestry samples, we detected 18 potentially novel BP loci (P< 5.0 x 10(-8)) in Stage 1 that warrant further replication. Additionally, correlated meta-analysis identified eight novel BP loci (11 genes). Several genes in these loci (e.g., PINX1, GATA4, BLK, FTO and GABBR2 have been previously reported to be associated with alcohol consumption. These findings provide insights into the role of alcohol consumption in the genetic architecture of hypertension

Pore size of macroporous polystyrene microspheres affects lipase immobilization

Polystyrene (PST) microspheres are commonly chosen as immobilization carriers due to their unique advantages such as ideal mechanical strength, adjustable particle size, and favorable chemical stability. However, there were few reports on immobilization by using microspheres with large pore sizes (>100 nm). We have successfully prepared the PST microspheres with macropores and gigapores by a novel method. In this study, giga-/macro-/meso-porous PST microspheres (314 nm, 104 nm, and 14.7 nm in pore sizes) were employed to immobilize lipase (from Burkholderia cepacia) by strong hydrophobic interactions, and the effects of pore sizes on lipase distribution, relative activity, kinetic behavior, thermal stability, storage stability, and reusability were also investigated in detail. According to laser scanning confocal microscope (LSCM) observation, lipase penetrated into the center of those giga-/macro-porous microspheres. With regard to the mesoporous microspheres, lipase only adsorbed to the external shell. The relative activities of immobilized lipase were 146%, 126%, and 50.9% for giga-/macro-/meso-porous PST microspheres, respectively. Comparing with the kinetic constants of free lipase (0.441 mM), the K(m) value for mesoporous PST-lipase (0.532 mM) was higher, whereas for giga-/macro-porous PST-lipase (0.402 mM and 0.411 mM), the K(m) values were comparatively lower suggesting the accessibility of substrate to the enzyme active sites was unlimited. The thermal stability, storage stability, and reusability were all improved significantly with the increase of pore sizes. In stimulant system, even after 100 times of recycling, the activity of lipase immobilized on gigaporous and macroporous PST microspheres remained nearly 100% and 93%, respectively, while that of lipase-PST with 14.7 nm pores could only keep 64.1%. In real system, 73.5% and 68.8% activity of gigaporous PST-lipase and macroporous PST-lipase, respectively, were retained after being used 30 times, whereas only 49.4% activity was remained for mesoporous PST-lipase. Therefore, compared with other microspheres, the gigaporous PST microsphere exhibited obvious advantages as a potential enzyme support in industry. (C) 2010 Elsevier B.V. All rights reserved

Enhanced induction of microspore embryogenesis after n-butanol treatment in wheat (Triticum aestivum L.) anther culture

The original version is available at http://www.springerlink.com/content/100383/The aim of this study was the improvement of embryo production in wheat anther culture. Three butanol alcohols, n-butanol, sec-butanol and tert-butanol, were evaluated for their effect on microspore embryogenesis in two spring cultivars of wheat, Pavon and Caramba. Application of n-butanol, at 0.1 and 0.2% (v/v) in the induction media for five hours, highly improved embryo production in both cultivars. Sec- and tert-butanol performed similarly to control plates. Regeneration ability was unaffected by any butyl-alcohol treatment. As a consequence of the higher embryo production after n-butanol treatment, the number of green regenerated plants increased up to 5 times in cultivar Pavon and up to 3 times in cultivar Caramba. The percentage of green plants was improved or unaffected by the treatment. Doubled haploid plant production was between 2 and 4 times higher after n-butanol treatment than in control plates. Therefore, n-butanol was successfully applied in the production of wheat doubled haploids. This primary alcohol is known as an activator of phospholipase D and has been previously reported to disrupt cortical microtubules and detach them from the plasma membrane in plants. Its effects on androgenetic induction could confirm the importance of microtubule regulation in plant cell fate, specifically in microspore development. A possible implication of phospholipase D is discussed.http://www.springerlink.com/content/v302947634352057/fulltext.pdfPeer reviewe