Modulatory effect of cAMP on fungal ergosterol level and inhibitory activity of azole drugs

The functions and biosynthesis of sterols have been effective targets for fungal control in different areas, including pharmaceutical and agricultural applications. Fungi are among the organisms that synthesize sterols, principally ergosterol. In this paper, the effect of dibutyryl-cAMP (db-cAMP) on ergosterol level and the interaction of drugs that would change the concentration of cAMP with antifungal drugs have been investigated. Sterols were extracted from Candida albicans, and ergosterol was measured using the gas chromatography method. The interaction of different agents was measured by the broth dilution method. It was found that phosphodiesterase inhibitors reverse the inhibitory activity of azole antifungal drugs. Evaluating the ergosterol level of C. albicans incubated with db-cAMP revealed that it increased ergosterol level. Further experiments provided evidence attributing the observed interaction between azoles and phosphodiesterase inhibitors to the relationship between ergosterol and cAMP. The possible significance of this interaction includes potentiation of antifungal activity of drugs by manipulating the cAMP level.     

AurH1: a new heptapeptide derived from Aurein1.2 antimicrobial peptide with specific and exclusive fungicidal activity

Due to the increasing incidence of fungal opportunistic infections and emergence of antibiotic‐resistant fungal strains, antimicrobial peptides (AMPs) are considered as ideal candidates for antifungal compounds. In silico methods can reduce the limitations of natural AMPs such as toxicity and instability and improve their antimicrobial properties and selectivity. In this study, we designed AurH1, a new truncated peptide, based on the six‐amino acid sequence of Aurein1.2. Further , the antimicrobial activities and toxicity effects of AurH1 on human skin fibroblast cells and red blood cells were investigated. Finally, field emission scanning electron microscopy (FE‐SEM) and flow cytometry were performed in order to study the mechanism of action of AurH1. The results indicated that AurH1 had only antifungal activity (at a minimal inhibitory concentration (MIC) of 7.3‐125 μg/mL) without any antibacterial effects on the selected bacteria, while Aurein1.2 had both antifungal and antibacterial activities as positive control. Furthermore, AurH1 did not show any toxicity on Hu02 cells and human red blood cells at its MIC range. In conclusion, it became clear that AurH1 is a selective peptide against fungi with no toxic effects on the selected bacteria and human cells.     

Sequence-dependent off-target inhibition of TLR7/8 sensing by synthetic microRNA inhibitors

Anti-microRNA (miRNA) oligonucleotides (AMOs) with 2′-O-Methyl (2′OMe) residues are commonly used to study miRNA function and can achieve high potency, with low cytotoxicity. Not withstanding this, we demonstrate the sequence-dependent capacity of 2′OMe AMOs to inhibit Toll-like receptor (TLR) 7 and 8 sensing of immunostimulatory RNA, independent of their miRNA-targeting function. Through a screen of 29 AMOs targeting common miRNAs, we found a subset of sequences highly inhibitory to TLR7 sensing in mouse macrophages. Interspecies conservation of this inhibitory activity was confirmed on TLR7/8 activity in human peripheral blood mononuclear cells. Significantly, we identified a core motif governing the inhibitory activity of these AMOs, which is present in more than 50 AMOs targeted to human miRNAs in miRBaseV20. DNA/locked nucleic acids (LNA) AMOs synthesized with a phosphorothioate backbone also inhibited TLR7 sensing in a sequence-dependent manner, demonstrating that the off-target effects of AMOs are not restricted to 2′OMe modification. Taken together, our work establishes the potential for off-target effects of AMOs on TLR7/8 function, which should be taken into account in their therapeutic development and in vivo application.     

Expression of a Novel Chimeric-Truncated tPA in Pichia pastoris with Improved Biochemical Properties

Thrombolytic therapy by plasminogen activators (PAs) has been a main goal in the treatment of acute myocardial infarction. Despite improved outcomes of currently available thrombolytic therapies, all these agents have different drawbacks that may result in less than optimal outcomes. In order to make tissue plasminogen activator (tPA) more potent, while being more resistant to plasminogen activator inhibitor-1 (PAI-1) and having a higher affinity to fibrin, a new chimeric-truncated form of tPA (CT tPA) was designed and expressed in Pichia pastoris. This novel variant consists of a finger domain of Desmoteplase, an epidermal growth factor (EGF) domain, a kringle 1 (K1) domain, a kringle 2 (K2) domain, in which the lysine binding site (LBS) was deleted, and a protease domain, where the four amino acids lysine 296, arginine 298, arginine 299, and arginine 304 were substituted by aspartic acid. The chimera CT tPA showed 14-fold increase in its activity in the presence of fibrin compared to the absence of fibrin. Furthermore, CT tPA showed about 10-fold more potency than commercially available full-length tPA (Actylase^®) and provided 1.2-fold greater affinity to fibrin. A residual activity of only 68 % was observed after incubation of Actylase^® with PAI-1, however, 91 % activity remained for CT tPA. These promising findings suggest that the novel CT tPA variant might be an acceptable PA with superior characteristics and properties.     

Automatic detection and resolution of measurement-unit conflicts in aggregated data

Background

Measurement-unit conflicts are a perennial problem in integrative research domains such as clinical meta-analysis. As multi-national collaborations grow, as new measurement instruments appear, and as Linked Open Data infrastructures become increasingly pervasive, the number of such conflicts will similarly increase.

Methods

We propose a generic approach to the problem of (a) encoding measurement units in datasets in a machine-readable manner, (b) detecting when a dataset contained mixtures of measurement units, and (c) automatically converting any conflicting units into a desired unit, as defined for a given study.

Results

We utilized existing ontologies and standards for scientific data representation, measurement unit definition, and data manipulation to build a simple and flexible Semantic Web Service-based approach to measurement-unit harmonization. A cardiovascular patient cohort in which clinical measurements were recorded in a number of different units (e.g., mmHg and cmHg for blood pressure) was automatically classified into a number of clinical phenotypes, semantically defined using different measurement units.

Conclusions

We demonstrate that through a combination of semantic standards and frameworks, unit integration problems can be automatically detected and resolved.

     

MicroRNA-15 family targets the hedgehog signaling pathway during postnatal development of porcine intestine

The hedgehog signaling pathway is a crucial regulator of the postnatal intestinal development. Regulation of hedgehog expression itself is poorly understood. MicroRNAs were demonstrated to control differentiation and proliferation in postnatal intestinal development. This study identifies members of the miR-15 family to regulate the expression of key hedgehog factors employing in silico and in vitro experiments. Physiological relevance is demonstrated by incorporation of in vivo expression data from the ileum and colon from 7 to 56 days old piglets. Results presented in this study improve the understanding of the complex regulation of hedgehog signaling during intestinal development and disease.     

Coenzyme Q10 Alleviated Behavioral Dysfunction and Bioenergetic Function in an Animal Model of Depression

Neurochemical Research - Coenzyme Q10 (CoQ10) is a natural compound, is involved in the mitochondrial electron transfer chain (ETC) and plays an important pattern in adenosine triphosphate (ATP)...     

La-related Protein 1 (LARP1) Represses Terminal Oligopyrimidine (TOP) mRNA Translation Downstream of mTOR Complex 1 (mTORC1)

The mammalian target of rapamycin complex 1 (mTORC1) is a critical regulator of protein synthesis. The best studied targets of mTORC1 in translation are the eukaryotic initiation factor-binding protein 1 (4E-BP1) and ribosomal protein S6 kinase 1 (S6K1). In this study, we identify the La-related protein 1 (LARP1) as a key novel target of mTORC1 with a fundamental role in terminal oligopyrimidine (TOP) mRNA translation. Recent genome-wide studies indicate that TOP and TOP-like mRNAs compose a large portion of the mTORC1 translatome, but the mechanism by which mTORC1 controls TOP mRNA translation is incompletely understood. Here, we report that LARP1 functions as a key repressor of TOP mRNA translation downstream of mTORC1. Our data show the following: (i) LARP1 associates with mTORC1 via RAPTOR; (ii) LARP1 interacts with TOP mRNAs in an mTORC1-dependent manner; (iii) LARP1 binds the 5′TOP motif to repress TOP mRNA translation; and (iv) LARP1 competes with the eukaryotic initiation factor (eIF) 4G for TOP mRNA binding. Importantly, from a drug resistance standpoint, our data also show that reducing LARP1 protein levels by RNA interference attenuates the inhibitory effect of rapamycin, Torin1, and amino acid deprivation on TOP mRNA translation. Collectively, our findings demonstrate that LARP1 functions as an important repressor of TOP mRNA translation downstream of mTORC1.     

Petal: a reliable explant for direct bulblet regeneration of endangered wild populations of <Emphasis Type="Italic">Fritillaria imperialis</Emphasis> L.

Wild populations of Fritillaria imperialis L. are facing extinction and need urgent conservation. This paper presents an efficient system for in vitro direct bulblet regeneration of these populations by petal culturing of flower buds. Petals at different developmental stages, green-closed flower bud (before nectar secretion) and red-closed flower bud (beginning of nectar secretion), were used as explants, and the effects of various proportions of cytokinin to auxin on direct bulblet regeneration pathway were evaluated. More explants switched on direct regeneration pathway in combination of auxins (0.6 mg l⁻¹ NAA + 0.4 mg l⁻¹ IAA) with higher level of cytokinin (1 mg l⁻¹ BAP). In contrast, auxins (0.6 mg l⁻¹ NAA + 0.4 mg l⁻¹ IAA) with lower level of cytokinin (0.1 mg l⁻¹ BAP) produced more bulblets per regenerated explant. In green-closed flower bud stage, direct bulblets regenerated from the end of petal where it was connected to the receptacle, while nectar secretion site was the place of bulblet formation in red-closed flower bud stage. In addition, genotype-dependency of direct bulblet regeneration pathway was investigated by using two different wild populations of Fritillaria imperialis. This plant regeneration procedure was applicable to different Fritillaria genotypes and regenerated bulblets were normal.     

Arginine-Rich Motifs Are Not Required for Hepatitis Delta Virus RNA Binding Activity of the Hepatitis Delta Antigen

Hepatitis delta virus (HDV) replication and packaging require interactions between the unbranched rodlike structure of HDV RNA and hepatitis delta antigen (HDAg), a basic, disordered, oligomeric protein. The tendency of the protein to bind nonspecifically to nucleic acids has impeded analysis of HDV RNA protein complexes and conclusive determination of the regions of HDAg involved in RNA binding. The most widely cited model suggests that RNA binding involves two proposed arginine-rich motifs (ARMs I and II) in the middle of HDAg. However, other studies have questioned the roles of the ARMs. Here, binding activity was analyzed in vitro using HDAg-160, a C-terminal truncation that binds with high affinity and specificity to HDV RNA segments in vitro. Mutation of the core arginines of ARM I or ARM II in HDAg-160 did not diminish binding to HDV unbranched rodlike RNA. These same mutations did not abolish the ability of full-length HDAg to inhibit HDV RNA editing in cells, an activity that involves RNA binding. Moreover, only the N-terminal region of the protein, which does not contain the ARMs, was cross-linked to a bound HDV RNA segment in vitro. These results indicate that the amino-terminal region of HDAg is in close contact with the RNA and that the proposed ARMs are not required for binding HDV RNA. Binding was not reduced by mutation of additional clusters of basic amino acids. This result is consistent with an RNA-protein complex that is formed via numerous contacts between the RNA and each HDAg monomer.