Characterization of pACK4, a mobilizable plasmid from Staphylococcus simulans biovar staphylolyticus

Staphylococcus simulans biovar staphylolyticus, the lysostaphin-producing organism, contains five plasmids designated pACK1–pACK5. pACK4 was found to be relaxable and to share sequence similarity with a number of well-characterized mobilizable plasmids from other staphylococci. All mobilizable staphylococcal plasmids characterized to date mediate resistance to various antibiotics, but pACK4 is unique because it contains no recognizable antibiotic resistance genes. pACK4 was found to contain an origin of transfer (oriT) region that shares inverted repeat regions and the same nic site as several other mobilizable staphylococcal plasmids. The presence of this conserved oriT region suggested that pACK4 might be mobilized in the presence of a conjugative plasmid. Filter mating studies revealed that pACK4 was mobilized by the conjugative plasmid pGO1. In addition, pACK4 was found to be virtually identical to the recently described plasmid pVGA from Staphylococcus aureus, except that pVGA contains an additional region (vgaA) that confers resistance to pleuromutilin, streptogramin A, and lincosamide. The high sequence similarity among pACK4, pVGA, and several previously described mobilizable staphylococcal plasmids suggests a common origin for these plasmids.     

Pyridopyrimidine based cannabinoid-1 receptor inverse agonists: Synthesis and biological evaluation

The synthesis, SAR and binding affinities are described for cannabinoid-1 receptor (CB1R) specific inverse agonists based on pyridopyrimidine and heterotricyclic scaffolds. Food intake and pharmacokinetic evaluation of several of these compounds indicate that they are effective orally active modulators of CB1R.     

Liberal and Conservative Protestant Denominations as Different Socioecological Strategies

It is common to portray conservative and liberal Protestant denominations as “strong” and “weak” on the basis of indices such as church attendance. Alternatively, they can be regarded as qualitatively different cultural systems that coexist in a multiple-niche environment. We integrate these two perspectives with a study of American teenagers based on both one-time survey information and the experience sampling method (ESM), which records individual experience on a moment-by-moment basis. Conservative Protestant youth were found to be more satisfied, family-oriented, and sociable than liberal Protestant youth, but also more dependent on their social environment, which is reflected in a deterioration of their mood when they are alone. Liberal Protestant youth appear to have internalized values that remain constant whether in the presence or absence of others. We relate these results to the social scientific literature on liberalism and conservatism and to evolutionary theory as a framework for explaining cultural systems as adaptations to multiple-niche environments.

QMEANclust: estimation of protein model quality by combining a composite scoring function with structural density information

Background  

The selection of the most accurate protein model from a set of alternatives is a crucial step in protein structure prediction both in template-based and ab initio approaches. Scoring functions have been developed which can either return a quality estimate for a single model or derive a score from the information contained in the ensemble of models for a given sequence. Local structural features occurring more frequently in the ensemble have a greater probability of being correct. Within the context of the CASP experiment, these so called consensus methods have been shown to perform considerably better in selecting good candidate models, but tend to fail if the best models are far from the dominant structural cluster. In this paper we show that model selection can be improved if both approaches are combined by pre-filtering the models used during the calculation of the structural consensus.     

Modulation of protein tyrosine phosphorylation in gastric mucosa during re-epithelization processes

AIM: To investigate the role of protein tyrosine phosphorylation in gastric wound formation and repair following ulceration.METHODS: Gastric lesions were induced in rats using restraint cold stress. To investigate the effect of oxidative and nitrosative cell stress on tyrosine phosphorylation during wound repair, total activity of protein tyrosine kinase (PTK), protein tyrosine phosphatase (PTP), antioxidant enzymes, nitric oxide synthase (NOS), 2’,5’-oligoadenylate synthetase, hydroxyl radical and zinc levels were assayed in parallel.RESULTS: Ulcer provocation induced an immediate decrease in tyrosine kinase (40% in plasma membranes and 56% in cytosol, P < 0.05) and phosphatase activity (threefold in plasma membranes and 3.3-fold in cytosol), followed by 2.3-2.4-fold decrease (P < 0.05) in protein phosphotyrosine content in the gastric mucosa. Ulceration induced no immediate change in superoxide dismutase (SOD) activity, 30% increase (P < 0.05) in catalase activity, 2.3-fold inhibition (P < 0.05) of glutathione peroxidase, 3.3-fold increase (P < 0.05) in hydroxyl radical content, and 2.3-fold decrease (P < 0.05) in zinc level in gastric mucosa. NOS activity was three times higher in gastric mucosa cells after cold stress. Following ulceration, PTK activity increased in plasma membranes and reached a maximum on day 4 after stress (twofold increase, P < 0.05), but remained inhibited (1.6-3-fold decrease on days 3, 4 and 5, P < 0.05) in the cytosol. Tyrosine phosphatases remained inhibited both in membranes and cytosol (1.5-2.4-fold, P < 0.05). NOS activity remained increased on days 1, 2 and 3 (3.8-, 2.6-, 2.2-fold, respectively, P < 0.05). Activity of SOD increased 1.6 times (P < 0.05) days 4 and 5 after stress. Catalase activity normalized after day 2. Glutathione peroxidase activity and zinc level decreased (3.3- and 2-fold, respectively, P < 0.05) on the last day. Activity of 2’,5’-oligoadenylate synthethase increased 2.8-fold (P < 0.05) at the beginning, and 1.6-2.3-fold (P < 0.05) during ulcer recuperation, and normalized on day 5, consistent with slowing of inflammation processes.CONCLUSION: These studies show diverse changes in total tyrosine kinase activity in gastric mucosa during the recovery process. Oxidative and nitrosative stress during lesion formation might lead to the observed reduction in tyrosine phosphorylation during ulceration.     

Factors influencing in the response of <i>Schizolobium parahybum</i> (Vell) Blake to <i>Ceratocystis paradoxa</i> and <i>C. moniliformis</i>

In the Ecuadorian coast one of the most destructive diseases of the pachaco is vascular wilt or stem rot caused by Ceratocystis complex, so the aim of this study was to determine the factors that affect the efficiency of the reaction of bark pachaco to this disease. This research was conducted under laboratory conditions, using trees pachaco S38, S41, S98, AE-1, AE-2 and AE-3, and pathogenic species Ceratocystis paradoxa and C. moniliformis. The method utilized was tissue stem bark,with bark sections with 4.5 cm², and a suspension of 3x10⁴ units infection and remained in a humid chamber for 96 hours at 25 ± 5 °C. Were determined grades of resistance/ susceptibility using a scale from 0 to 4, depending on the amount of mycelia and peritecio in each plant sample. Three factors were used: four colonies obtained by several transfers from each fungal specie, four ages of colonies of each fungal specie and four volumes of inoculum applied (units of infection), using for each experiment separately Completely Randomized Design with 4 replications factorial arrangement. For comparison between treatment means was used Tukey test at 5% probability of error. For future trials using this technique, you could use 30-day colonies for C. paradoxa and 40 days for C. moniliformis, and an application volume of 100 μL/cm², it would improve the level of response for the formation of perithecium and mycelia in samples cortex.     

Second-site compensatory mutations of HIV-1 capsid mutations

The human immunodeficiency virus (HIV) capsid (CA) protein assembles into a hexameric lattice that forms the mature virus core. Contacts between the CA N-terminal domain (NTD) of one monomer and the C-terminal domain (CTD) of the adjacent monomer are important for the assembly of this core. In this study, we have examined the effects of mutations in the NTD region associated with this interaction. We have found that such mutations yielded modest reductions of virus release but major effects on viral infectivity. Cell culture and in vitro assays indicate that the infectivity defects relate to abnormalities in the viral cores. We have selected second-site compensatory mutations that partially restored HIV infectivity. These mutations map to the CA CTD and to spacer peptide 1 (SP1), the portion of the precursor Gag protein immediately C terminal to the CTD. The compensatory mutations do not locate to the molecularly modeled intermolecular NTD-CTD interface. Rather, the compensatory mutations appear to act indirectly, possibly by realignment of the C-terminal helix of the CA CTD, which participates in the NTD-CTD interface and has been shown to serve an important role in the assembly of infectious virus.     

Histone Deacetylase Inhibitor Romidepsin Induces HIV Expression in CD4 T Cells from Patients on Suppressive Antiretroviral Therapy at Concentrations Achieved by Clinical Dosing

Persistent latent reservoir of replication-competent proviruses in memory CD4 T cells is a major obstacle to curing HIV infection. Pharmacological activation of HIV expression in latently infected cells is being explored as one of the strategies to deplete the latent HIV reservoir. In this study, we characterized the ability of romidepsin (RMD), a histone deacetylase inhibitor approved for the treatment of T-cell lymphomas, to activate the expression of latent HIV. In an in vitro T-cell model of HIV latency, RMD was the most potent inducer of HIV (EC₅₀ = 4.5 nM) compared with vorinostat (VOR; EC₅₀ = 3,950 nM) and other histone deacetylase (HDAC) inhibitors in clinical development including panobinostat (PNB; EC₅₀ = 10 nM). The HIV induction potencies of RMD, VOR, and PNB paralleled their inhibitory activities against multiple human HDAC isoenzymes. In both resting and memory CD4 T cells isolated from HIV-infected patients on suppressive combination antiretroviral therapy (cART), a 4-hour exposure to 40 nM RMD induced a mean 6-fold increase in intracellular HIV RNA levels, whereas a 24-hour treatment with 1 µM VOR resulted in 2- to 3-fold increases. RMD-induced intracellular HIV RNA expression persisted for 48 hours and correlated with sustained inhibition of cell-associated HDAC activity. By comparison, the induction of HIV RNA by VOR and PNB was transient and diminished after 24 hours. RMD also increased levels of extracellular HIV RNA and virions from both memory and resting CD4 T-cell cultures. The activation of HIV expression was observed at RMD concentrations below the drug plasma levels achieved by doses used in patients treated for T-cell lymphomas. In conclusion, RMD induces HIV expression ex vivo at concentrations that can be achieved clinically, indicating that the drug may reactivate latent HIV in patients on suppressive cART.