Exploring QSAR of thiazole and thiadiazole derivatives as potent and selective human adenosine A3 receptor antagonists using FA and GFA techniques

Binding affinity data of thiazole and thiadiazole derivatives (n=30) for human adenosine A3 receptor subtype have been subjected to Quantitative Structure-Activity Relationship (QSAR) analysis using quantum chemical and hydrophobicity parameters. Wang-Ford charges of the common atoms of the compounds [calculated from molecular electrostatic potential surface of energy minimized geometry using Austin Model 1 (AM1) technique] were used as independent variables apart from partition coefficient (logP) and suitable dummy parameters. The variables for the multiple regression analyses were selected based on principal component factor analysis (FA), and generated equations were statistically validated using leave-one-out technique. The best equation thus obtained explained and predicted 74.4% and 68.9% respectively of the variance of the binding affinity. The results suggested importance of Wang-Ford charges of atoms C2, C5 and C7. Furthermore, the A3 binding affinity increases with decrease of lipophilicity of the compounds and in the presence of methyl or ethyl substituent at R position. Again, the binding affinity decreases in the presence of tert-butyloxy group at R position. When factor scores were used as predictor variables in principal component regression analysis, the resulted model showed 87.0% predicted variance and 89.5% explained variance. The data set was also modeled using genetic function approximation (GFA) technique. The best two equations derived from GFA show better predicted variance values (0.753 and 0.739) than that found in case of the best equation derived from FA. However, considerable intercorrelation was found between two predictor variables in case of GFA derived equations. GFA derived equations show importance of Wang-Ford charges of different atoms of the thiazole/thiadiazole nucleus and phenyl ring (S9, X8 and C2, the effects of the first two being predominant) along with similar impact of lipophilicity and R group on the binding affinity as found in case of the FA derived relation.     

Improved understanding of gene expression regulation using systems biology

This article reviews the current state of systems biology approaches, including the experimental tools used to generate 'omic' data and computational frameworks to interpret this data. Through illustrative examples, systems biology approaches to understand gene expression and gene expression regulation are discussed. Some of the challenges facing this field and the future opportunities in the systems biology era are highlighted.     

Benchmarking of 16S rRNA gene databases using known strain sequences

16S rRNA gene analysis is the most convenient and robust method for microbiome studies. Inaccurate taxonomic assignment of bacterial strains could have deleterious effects as all downstream analyses rely heavily on the accurate assessment of microbial taxonomy. The use of mock communities to check the reliability of the results has been suggested. However, often the mock communities used in most of the studies represent only a small fraction of taxa and are used mostly as validation of sequencing run to estimate sequencing artifacts. Moreover, a large number of databases and tools available for classification and taxonomic assignment of the 16S rRNA gene make it challenging to select the best-suited method for a particular dataset. In the present study, we used authentic and validly published 16S rRNA gene type strain sequences (full length, V3-V4 region) and analyzed them using a widely used QIIME pipeline along with different parameters of OTU clustering and QIIME compatible databases. Data Analysis Measures (DAM) revealed a high discrepancy in ratifying the taxonomy at different taxonomic hierarchies. Beta diversity analysis showed clear segregation of different DAMs. Limited differences were observed in reference data set analysis using partial (V3-V4) and full-length 16S rRNA gene sequences, which signify the reliability of partial 16S rRNA gene sequences in microbiome studies. Our analysis also highlights common discrepancies observed at various taxonomic levels using various methods and databases.     

Genetic Analysis of Circadian Responses to Low Frequency Electromagnetic Fields in Drosophila melanogaster

The blue-light sensitive photoreceptor cryptochrome (CRY) may act as a magneto-receptor through formation of radical pairs involving a triad of tryptophans. Previous genetic analyses of behavioral responses of Drosophila to electromagnetic fields using conditioning, circadian and geotaxis assays have lent some support to the radical pair model (RPM). Here, we describe a new method that generates consistent and reliable circadian responses to electromagnetic fields that differ substantially from those already reported. We used the Schuderer apparatus to isolate Drosophila from local environmental variables, and observe extremely low frequency (3 to 50 Hz) field-induced changes in two locomotor phenotypes, circadian period and activity levels. These field-induced phenotypes are CRY- and blue-light dependent, and are correlated with enhanced CRY stability. Mutational analysis of the terminal tryptophan of the triad hypothesised to be indispensable to the electron transfer required by the RPM reveals that this residue is not necessary for field responses. We observe that deletion of the CRY C-terminus dramatically attenuates the EMF-induced period changes, whereas the N-terminus underlies the hyperactivity. Most strikingly, an isolated CRY C-terminus that does not encode the Tryptophan triad nor the FAD binding domain is nevertheless able to mediate a modest EMF-induced period change. Finally, we observe that hCRY2, but not hCRY1, transformants can detect EMFs, suggesting that hCRY2 is blue light-responsive. In contrast, when we examined circadian molecular cycles in wild-type mouse suprachiasmatic nuclei slices under blue light, there was no field effect. Our results are therefore not consistent with the classical Trp triad-mediated RPM and suggest that CRYs act as blue-light/EMF sensors depending on trans-acting factors that are present in particular cellular environments.     

Mitochondrial DNA analysis reveals three stocks of yellowfin tuna Thunnus albacares (Bonnaterre, 1788) in Indian waters

Yellowfin tuna (Thunnus albacares) is an epipelagic, oceanic species of family Scombridae found in tropical and subtropical region of Pacific, Indian and Atlantic Ocean. It is commercially important fish and accounts for 19 % of total tuna catches in Indian waters. In present study, population structure of yellowfin tuna was examined using sequence analysis of mitochondrial DNA from seven geographically distinct locations along the Indian coast. A 500 bp segment of D-loop region was sequenced and analysed for 321 yellowfin samples. Hierarchical analysis of molecular variance showed significant genetic differentiation among three groups (VE); (AG); (KO, TU, PO, VI, PB) analyzed (Φ _ST  = 0.03844, P ≤ 0.001). In addition, spatial analysis of molecular variance identified three genetically heterogeneous groups of yellowfin tuna in Indian waters. Results were further corroborated by significant value of nearest neighbour statistic (S _nn = 0.261, P ≤ 0.001). Thus finding of this study rejects the null hypothesis of single panmictic population of yellowfin tuna in Indian waters.     

In vitro conservation and asymbiotic propagation of Coelogyne flaccida (Lindl.): A threatened orchid

Asymbiotic seed germination of Coelogyne flaccida varied with the capsule stage and the culture medium used for germinating seeds. The capsules were harvested at two different stages of development. The seeds were cultured on three asymbiotic orchid seed germination defined and undefined media, i.e. Mitra (M) medium, Murashige and Skoog (MS) medium and potato dextrose agar (PDA) medium. The seeds obtained from undehisced green capsules germinated with a maximum germination percentage (84.50 ± 0.33%) on M medium followed by MS and PDA medium. The effect of cytokinins, such as 6-benzylaminopurine and furfurylaminopurine and the synthetic auxin α-naphthalene acetic acid, on seed germination was also assessed. Simultaneously, in vitro multiplication using protocorms as explants was also studied. The effect of organic growth supplements, such as banana homogenate (BH, 25, 50, 75 g l^? 1) and peptone (P, 1.0, 1.5, 2.0 g l^? 1), was tested on the de novo formation of protocorm-like bodies (PLBs), development of the maximum number of shoots and early formation of plantlets using the M medium. Among the treatments, the highest regeneration frequency (87.50 ± 0.20%) and the highest number of PLBs per explant (10.25 ± 0.50) were obtained in P (1.5 g l^? 1)-supplemented cultures, and the plantlets were formed within 18 weeks of culture. BH favoured the development of healthy plantlets, with a maximum fresh weight of 1.02 ± 0.04 g per plantlet.     

Pseudouridine formation in archaeal RNAs: The case of Haloferax volcanii

Pseudouridine (Ψ), the isomer of uridine, is commonly found at various positions of noncoding RNAs of all organisms. Ψ residues are formed by a number of single- or multisite specific Ψ synthases, which generally act as stand-alone proteins. In addition, in Eukarya and Archaea, specific ribonucleoprotein complexes, each containing a distinct box H/ACA guide RNA and four core proteins, can produce Ψ at many sites of different cellular RNAs. Cbf5 is the core Ψ synthase in these complexes. Using Haloferax volcanii as an archaeal model organism, we show that, contrary to eukaryotes, the Cbf5 homolog (HVO_2493) is not essential in this archaeon. The Cbf5-deleted strain of H. volcanii completely lacks Ψ at positions 1940, 1942, 2605, and 2591 (Escherichia coli positions 1915, 1917, 2572, and 2586) of its 23S rRNA, and contains reduced steady-state levels of some box H/ACA RNAs. Archaeal Cbf5 is known to have tRNA Ψ55 synthase activity in vitro but we could not confirm this activity in vivo in H. volcanii. Conversely, the Pus10 (previously PsuX) homolog (HVO_1979), which can produce tRNA Ψ55, as well as Ψ54 in vitro, is shown here to be essential in H. volcanii, whereas the corresponding tRNA Ψ55 synthases, Pus4 and TruB, are not essential in yeast and E. coli, respectively. Finally, we demonstrate that HVO_1852, the TruA/Pus3 homolog, is responsible for the pseudouridylation of position 39 in H. volcanii tRNAs and that the corresponding gene is not essential.     

Calcium signals and calpain-dependent necrosis are essential for release of coxsackievirus B from polarized intestinal epithelial cells

Coxsackievirus B (CVB), a member of the enterovirus family, targets the polarized epithelial cells lining the intestinal tract early in infection. Although the polarized epithelium functions as a protective barrier, this barrier is likely exploited by CVB to promote viral entry and subsequent egress. Here we show that, in contrast to nonpolarized cells, CVB-infected polarized intestinal Caco-2 cells undergo nonapoptotic necrotic cell death triggered by inositol 1,4,5-trisphosphate receptor-dependent calcium release. We further show that CVB-induced cellular necrosis depends on the Ca(2+)-activated protease calpain-2 and that this protease is involved in CVB-induced disruption of the junctional complex and rearrangements of the actin cytoskeleton. Our study illustrates the cell signaling pathways hijacked by CVB, and perhaps other viral pathogens, to promote their replication and spread in polarized cell types.