Genetic Characterization of Vibrio vulnificus Strains from Tilapia Aquaculture in Bangladesh

Outbreaks of Vibrio vulnificus wound infections in Israel were previously attributed to tilapia aquaculture. In this study, V. vulnificus was frequently isolated from coastal but not freshwater aquaculture in Bangladesh. Phylogenetic analyses showed that strains from Bangladesh differed remarkably from isolates commonly recovered elsewhere from fish or oysters and were more closely related to strains of clinical origin.Vibrio vulnificus causes severe wound infections and life-threatening septicemia (mortality, >50%), primarily in patients with underlying chronic diseases (10, 19, 23) and primarily from raw oyster consumption (21). This Gram-negative halophile is readily recovered from oysters (27, 35, 43) and fish (14) and was initially classified into two biotypes (BTs) based on growth characteristics and serology (5, 18, 39). Most human isolates are BT1, while BT2 is usually associated with diseased eels (1, 39). An outbreak of wound infections from aquacultured tilapia in Israel (6) revealed a new biotype (BT3). Phenotypic assays do not consistently distinguish biotypes (33), but genetic analyses have helped resolve relationships (20). A 10-locus multilocus sequence typing (MLST) scheme (8, 9) and a similar analysis of 6 loci (13) segregated V. vulnificus strains into two clusters. BT1 strains were in both clusters, while BT2 segregated into a single cluster and BT3 was a genetic mosaic of the two lineages. Significant associations were observed between MLST clusters and strain origin: most clinical strains (BT1) were in one cluster, and the other cluster was comprised mostly of environmental strains (some BT1 and all BT2). Clinical isolates were also associated with a unique genomic island (13).The relationship between genetic lineages and virulence has not been determined, and confirmed virulence genes are universally present in V. vulnificus strains from both clinical and environmental origins (19, 23). However, segregation of several polymorphic alleles agreed with the MLST analysis and correlated genotype with either clinical or environmental strain origin. Alleles include 16S rRNA loci (15, 26, 42), a virulence-correlated gene (vcg) locus (31, 41, 42), and repetitive sequence in the CPS operon (12). DiversiLab repetitive extrageneic palindromic (rep-PCR) analysis also confirmed these genetic distinctions and showed greater diversity among clinical strains (12).Wound infections associated with tilapia in Israel implicated aquaculture as a potential source of V. vulnificus in human disease (6, 40). Tilapia aquaculture is increasing rapidly, as shown by a 2.8-fold increase in tons produced from 1998 to 2007 (Food and Agriculture Organization; http://www.fao.org/fishery/statistics/en). Therefore, presence of V. vulnificus in tilapia aquaculture was examined in Bangladesh, a region that supports both coastal and freshwater sources of industrial-scale aquaculture. V. vulnificus strains were recovered from market fish, netted fish, and water samples, and the phylogenetic relationship among strains was examined relative to clinical and environmental reference strains collected elsewhere.     

人HCN4通道的滞后现象:影响窦房结起搏的潜在决定因素(英文)

超极化活化环核苷酸门控(hyperpolarization-activated cyclic-nucleotide-gated,HCN)通道参与调制心脏跳动的节律和速率。与HCN1和HCN2有所不同,慢通道HCN4可能不存在电压依赖的滞后现象。本研究采用单细胞膜片钳方法,在稳定转染hHCN4的HEK293细胞上进行电生理记录,观察hHCN4通道是否存在滞后现象,以及cAMP对其的调制作用;同时采用实时定量RT-PCR方法检测窦房结和心房组织中HCNs的表达。电压钳实验结果显示hHCN4电流(Ih)激活随着保持电位超极化的变化而向去极化方向移动。三角电位变化钳(triangular ramp)和动作电位钳的结果也显示了hHCN4的滞后现象。cAMP增加Ih电流幅度,且使电流激活向去极化方向移动,从而改变内源性hHCN4滞后行为。RT-PCR结果显示,人窦房结组织主要表达HCN4,占75%,HCN1占21%,HCN2占3%,HCN3占0.7%。以上结果提示,人窦房结组织主要表达HCN4亚型,hHCN4的Ih存在电压依赖性的滞后现象,且受cAMP调制。由此推断,hHCN4通道的滞后现象可能在窦房结起搏活动中起到了关键作用。     

Benzene and dopamine catechol quinones could initiate cancer or neurogenic disease

Catechol quinones of estrogens react with DNA by 1,4-Michael addition to form depurinating N3Ade and N7Gua adducts. Loss of these adducts from DNA creates apurinic sites that can generate mutations leading to cancer initiation. We compared the reactions of the catechol quinones of the leukemogenic benzene (CAT-Q) and N-acetyldopamine (NADA-Q) with 2′-deoxyguanosine (dG) or DNA. NADA was used to prevent intramolecular cyclization of dopamine quinone. Reaction of CAT-Q or NADA-Q with dG at pH 4 afforded CAT-4-N7dG or NADA-6-N7dG, which lost deoxyribose with a half-life of 3 h to form CAT-4-N7Gua or 4 h to form NADA-6-N7Gua. When CAT-Q or NADA-Q was reacted with DNA, N3Ade adducts were formed and lost from DNA instantaneously, whereas N7Gua adducts were lost over several hours. The maximum yield of adducts in the reaction of CAT-Q or NADA-Q with DNA at pH 4 to 7 was at pH 4. When tyrosinase-activated CAT or NADA was reacted with DNA at pH 5 to 8, adduct levels were much higher (10- to 15-fold), and the highest yield was at pH 5. Reaction of catechol quinones of natural and synthetic estrogens, benzene, naphthalene, and dopamine with DNA to form depurinating adducts is a common feature that may lead to initiation of cancer or neurodegenerative disease.     

Structural basis of TRAPPIII‐mediated Rab1 activation

The GTPase Rab1 is a master regulator of the early secretory pathway and is critical for autophagy. Rab1 activation is controlled by its guanine nucleotide exchange factor, the multisubunit TRAPPIII complex. Here, we report the 3.7 Å cryo‐EM structure of the Saccharomyces cerevisiae TRAPPIII complex bound to its substrate Rab1/Ypt1. The structure reveals the binding site for the Rab1/Ypt1 hypervariable domain, leading to a model for how the complex interacts with membranes during the activation reaction. We determined that stable membrane binding by the TRAPPIII complex is required for robust activation of Rab1/Ypt1 in vitro and in vivo, and is mediated by a conserved amphipathic α‐helix within the regulatory Trs85 subunit. Our results show that the Trs85 subunit serves as a membrane anchor, via its amphipathic helix, for the entire TRAPPIII complex. These findings provide a structural understanding of Rab activation on organelle and vesicle membranes.     

DF++ : an adaptive buffer-aware probabilistic delegation forwarding protocol for Delay Tolerant Network

The delegated forwarding (DF) curb transmissions by forwarding the message to a node that holds high quality value seen by the message. However, DF assumes infinite buffer space that is not possible in real time applications. In addition, quality value computation considers the encountering history and does not account for additional network parameters such as aging and transitive connectivity. In this paper, we have proposed a routing protocol called as DF++ that compute quality value based on probabilistic model used in PRoPHET protocol and forwards the message to current node by adaptive computation of available buffer space. We have compared performance of DF++ with DF, Epidemic and PRoPHET routing protocols. The proposed DF++ has higher delivery probability and fewer message drop and transmissions.     

The modification of the peptidyl transferase activity of 50-S ribosomal subunits, LiC1-split proteins and L16 ribosomal protein by ethoxyformic anhydride.

Ethoxyformic anhydride abolishes the peptidyl transferase activity of 50-S ribosomal subunits, LiC1 split proteins and L16. Hydroxylamine treatment results in reactivation. Erythromycin exhibits significant protection with 50-S ribosomal subunits. With LiC1 split proteins and L16 significant protection was exhibited only after reconstitution. The results indicate that the ethoxyformic anhydride is reacting with approximately six histidines in LiC1 split proteins and one in L16. Since L16 has been reported to contain a single histidine, the results presented indicate the involvement of this histidine in peptidyl transferase activity.     

Lipase in aqueous‐polar organic solvents: Activity,structure, and stability

Studying alterations in biophysical and biochemical behavior of enzymes in the presence of organic solvents and the underlying cause(s) has important implications in biotechnology. We investigated the effects of aqueous solutions of polar organic solvents on ester hydrolytic activity, structure and stability of a lipase. Relative activity of the lipase monotonically decreased with increasing concentration of acetone, acetonitrile, and DMF but increased at lower concentrations (upto ～20% v/v) of dimethylsulfoxide, isopropanol, and methanol. None of the organic solvents caused any appreciable structural change as evident from circular dichorism and NMR studies, thus do not support any significant role of enzyme denaturation in activity change. Change in 2D [15N, 1H]‐HSQC chemical shifts suggested that all the organic solvents preferentially localize to a hydrophobic patch in the active‐site vicinity and no chemical shift perturbation was observed for residues present in protein's core. This suggests that activity alteration might be directly linked to change in active site environment only. All organic solvents decreased the apparent binding of substrate to the enzyme (increased K_m); however significantly enhanced the k_cat. Melting temperature (T_m) of lipase, measured by circular dichroism and differential scanning calorimetry, altered in all solvents, albeit to a variable extent. Interestingly, although the effect of all organic solvents on various properties on lipase is qualitatively similar, our study suggest that magnitudes of effects do not appear to follow bulk solvent properties like polarity and the solvent effects are apparently dictated by specific and local interactions of solvent molecule(s) with the protein.     

Prostate cancer is known by the companionship with ATM and miRNA it keeps: craftsmen of translation have dual behaviour with tailors of life thread

Research on prostate cancer progression has focused extensively on the concept of miRNA, which can operate either as promoters or as suppressors of carcinogenesis. Moreover, recent genetic studies and emerging functional work show that strikingly similar and overlapping pathways are involved in prostate carcinogenesis. Unswervingly, these elements constitute a recently explored ‘network of networks’ that dynamically reorganizes during DNA damage and is responsible for positively or negatively regulating genome organization and integrity. We consider these facets of convergence and discuss how insights from diametrically opposed interactions of ataxia–telangiectasia mutated and mitrons can inform us about, and possibly help us to get a step closer to personalized medicine. Copyright © 2012 John Wiley & Sons, Ltd.     

Platelet‐derived growth factor (PDGF) signalling in cancer: rapidly emerging signalling landscape

Platelet‐derived growth factor (PDGF)‐mediated signalling has emerged as one of the most extensively and deeply studied biological mechanism reported to be involved in regulation of growth and survival of different cell types. However, overwhelmingly increasing scientific evidence is also emphasizing on dysregulation of spatio‐temporally controlled PDGF‐induced signalling as a basis for cancer development. We partition this multi‐component review into recently developing understanding of dysregulation PDGF signalling in different cancers, how PDGF receptors are quantitatively controlled by microRNAs. Moreover, we also summarize most recent advancements in therapeutic targeting of PDGFR as evidenced by preclinical studies. Better understanding of the PDGF‐induced intracellular signalling in different cancers will be helpful in catalysing the transition from a segmented view of cancer biology to a conceptual continuum.