Cloning and characterization of an eukaryotic initiation factor-2alpha kinase from the silkworm,Bombyx mori

Eukaryotic initiation factor 2alpha (eIF-2alpha) kinases are involved in the translational regulations that occur in response to various types of environmental stress, and play an important role in the cellular defense system operating under unfavorable conditions. The identification of additional eIF-2alpha kinases and the elucidation of their functions are necessary to understand how different eIF-2alpha kinases can specifically respond to distinct stimuli. Here, we report a novel eIF-2alpha kinase, termed BeK, from the silkworm, Bombyx mori. This gene encodes 579 amino acids and contains all 11 catalytic domains of protein-serine/threonine kinases. Most notably, it contains an "Ile-Gln-Met-Xaa-Xaa-Cys" motif, which is highly conserved from yeast to mammalian eIF-2alpha kinases. BeK does not show any significant homology in the NH(2) terminal regulatory domain, suggesting a distinct regulatory mechanism of this novel eIF-2alpha kinase. BeK is ubiquitously expressed in the various tissues throughout the final larval stage. Importantly, BeK is activated in Drosophila Schneider cells following heat shock and osmotic stress, and activated-BeK has been shown to phosphorylate an eIF-2alpha subunit at the Ser(50) site. However, other forms of stress, such as immune stress, endoplasmic reticulum stress and oxidative stress, cannot significantly elicit BeK activity. Interestingly, the baculovirus gene product, PK2, can inhibit BeK enzymatic activity, suggesting that BeK may be an endogenous target for a viral gene product. Taken together, these data indicate that BeK is a novel eIF-2alpha kinase involved in the stress response in B. mori.     

Synthesis and biological evaluation of pyrazole linked benzothiazole-β-naphthol derivatives as topoisomerase I inhibitors with DNA binding ability

A series of new pyrazole linked benzothiazole-β-naphthol derivatives were designed and synthesized using a simple, efficient and ecofriendly route under catalyst-free conditions in good to excellent yields. These derivatives were evaluated for their cytotoxicity on selected human cancer cell lines. Among those, the derivatives 4j, 4k and 4l exhibited considerable cytotoxicity with IC₅₀ values ranging between 4.63 and 5.54?µM against human cervical cancer cells (HeLa). Structure activity relationship was elucidated by varying different substituents on benzothiazoles and pyrazoles. Further, flow cytometric analysis revealed that these derivatives induced cell cycle arrest in G2/M phase and spectroscopic studies such as UV–visible, fluorescence and circular dichroism studies showed that these derivatives exhibited good DNA binding affinity. Additionally, these derivatives can effectively inhibit the topoisomerase I activity. Viscosity studies and molecular docking studies demonstrated that the derivatives bind with the minor groove of the DNA.     

Distinct roles of chromatin-associated proteins MDC1 and 53BP1 in mammalian double-strand break repair

Phosphorylated histone H2AX ("gamma-H2AX") recruits MDC1, 53BP1, and BRCA1 to chromatin near a double-strand break (DSB) and facilitates efficient repair of the break. It is unclear to what extent gamma-H2AX-associated proteins act in concert and to what extent their functions within gamma-H2AX chromatin are distinct. We addressed this question by comparing the mechanisms of action of MDC1 and 53BP1 in DSB repair (DSBR). We find that MDC1 functions primarily in homologous recombination/sister chromatid recombination, in a manner strictly dependent upon its ability to interact with gamma-H2AX but, unexpectedly, not requiring recruitment of 53BP1 or BRCA1 to gamma-H2AX chromatin. In contrast, 53BP1 functions in XRCC4-dependent nonhomologous end-joining, likely mediated by its interaction with dimethylated lysine 20 of histone H4 but, surprisingly, independent of H2AX. These results suggest a specialized adaptation of the "histone code" in which distinct histone tail-protein interactions promote engagement of distinct DSBR pathways.     

Utilizing Waste Cable Wires for High‐Performance Fiber‐Based Hybrid Supercapacitors: An Effective Approach to Electronic‐Waste Management

In recent years, electronic waste (e‐waste) such as old cable wires, fans, circuit boards, etc., can be often seen in large piles of leftover in dumping yards. Employing these e‐waste sources for energy storage devices not only increases the economic value but also decreases the reliance on fossil fuels. In this context, waste cable wires are utilized to obtain precious copper (Cu) fibers and used as a cost‐effective current collector for the fabrication of fiber‐based hybrid supercapacitor (FHSC). With the braided Cu fibers, forest‐like nickel oxide nanosheet grafted carbon nanotube coupled copper oxide nanowire arrays (NiO NSs@CNTs@CuO NWAs/Cu fibers) are designed via simple wet‐chemical approaches. As a battery‐type material, the forest‐like NiO NSs@CNTs@CuO NWAs/Cu fiber electrode shows superior electrochemical properties including high specific capacity (230.48 mA h g^?1) and cycling stability (82.72%) in aqueous alkaline electrolyte. Moreover, a solid‐state FHSC is also fabricated using forest‐like NiO NSs@CNTs@CuO NWAs/Cu fibers as a positive electrode and activated carbon coated carbon fibers as a negative electrode with a gel electrolyte, which also shows a higher energy and power densities of 26.32 W h kg^?1 and 1218.33 W kg^?1, respectively. The flexible FHSC is further employed as an energy source for various electronic gadgets, demonstrating its suitability for wearable applications.     

Synthesis,computational studies and antiproliferative activities of coumarin-tagged 1,3,4-oxadiazole conjugates against MDA-MB-231 and MCF-7 human breast cancer cells

A novel library of coumarin tagged 1,3,4 oxadiazole conjugates was synthesized and evaluated for their antiproliferative activities against MDA-MB-231 and MCF-7 breast cancer cell lines. The evaluation studies revealed that compound 9d was the most potent molecule with an IC₅₀ value of <5?µM against the MCF-7 cell line. Interestingly, compounds 10b and 11a showed a similar trend with lower inhibitory concentration (IC₅₀?=?7.07?µM), in Estrogen Negative (ER?) cells than Estrogen Positive (ER+) cells. Structure–activity relationship (SAR) studies revealed that conjugates bearing benzyl moieties (9b, 9c and 9d) had superior activities compared to their alkyl analogues. The most potent compound 9d showed ～1.4?times more potent activity than tamoxifen against MCF-7 cell line; while the introduction of sulfone unit in compounds 11a, 11b and 11c resulted in significant cytotoxicity against both MCF-7 and MDA-MB-231 cell lines. These results were further supported by docking studies, which revealed that the stronger binding affinity of the synthesized conjugates is due to the presence of sulfone unit attached to the substituted benzyl moiety in their pharmacophores.     

Biochemical studies of amylases in the silkworm, Bombyx mori L.: Comparative analysis in diapausing and nondiapausing strains

Different amylase enzymes were identified by analysis of digestive fluid and haemolymph in diapausing and nondiapausing strains of silkworm, Bombyx mori. The diapausing strain showed negligible digestive amylase activity at a pH range of 3–11, while the nondiapausing strain registered strikingly higher amylase activity at pH 9.2. Higher levels of undigested starch was found in the faecal matter of the diapausing strain, which is consistent with the negligible digestive amylase activity. Development specific expression of haemolymph amylase activity was seen in nondiapausing and diapausing strains. In the nondiapausing strain the digestive amylase activity was at its peak during intermoult and depressed during moult. PAGE analysis revealed the occurrence of only anodal digestive and haemolymph amylases in the diapausing strain, whereas both cathodal and anodal enzymes were seen in the digestive fluid and haemolymph of the nondiapausing strain.     

Determination of albumin adducts of 4,4′-methylenediphenyl diisocyanate in workers of a 4,4′-methylenedianiline factory

Lung sensitization and asthma are the main health effects of 4,4′-methylenediphenyl diisocyanate (MDI). Albumin adducts (isocyanate specific adducts) of MDI might be involved in the etiology of sensitization reactions. Albumin adducts of MDI have been found in subjects classified as 4,4′-methylenedianiline (MDA) workers. The mean adduct levels in these MDA-workers were 1.5 times higher than in MDI-workers of the same company. MDA-specific hemoglobin adducts, were present ten times more in the MDA-workers than in the MDI-workers. MDA-workers with specific work task had significantly higher albumin adduct levels.     

Differences in the plasma membrane proteins of chronic alcoholic rat brain

Plasma membranes were isolated from the cerebral cortex of control and chronic ethanol-treated rat brains. Analysis of protein composition by SDS-PAGE and by two-dimensional gel electrophoresis (IEF-SDS-PAGE) revealed significant differences in the membrane protein patterns between control and ethanol-treated rat cerebral cortices, indicating the loss of several proteins in membranes from ethanol-treated rat brains. Plasma membrane-associated protein species are categorized into ethanol-sensitive and -insensitive proteins, based on their response to ethanol. This study reports that ethanol depletes certain intrinsic proteins of membranes that might be responsible for plasma membrane disruption by ethanol.     

Evaluation of Skeletal and Cardiac Muscle Function after Chronic Administration of Thymosin β-4 in the Dystrophin Deficient Mouse

Thymosin beta-4 (Tβ4) is a ubiquitous protein with many properties relating to cell proliferation and differentiation that promotes wound healing and modulates inflammatory mediators. We studied the effects of chronic administration of Tβ4 on the skeletal and cardiac muscle of dystrophin deficient mdx mice, the mouse model of Duchenne muscular dystrophy. Female wild type (C57BL10/ScSnJ) and mdx mice, 8–10 weeks old, were treated with 150 µg of Tβ4 twice a week for 6 months. To promote muscle pathology, mice were exercised for 30 minutes twice a week. Skeletal and cardiac muscle function were assessed via grip strength and high frequency echocardiography. Localization of Tβ4 and amount of fibrosis were quantified using immunohistochemistry and Gomori''s tri-chrome staining, respectively. Mdx mice treated with Tβ4 showed a significant increase in skeletal muscle regenerating fibers compared to untreated mdx mice. Tβ4 stained exclusively in the regenerating fibers of mdx mice. Although untreated mdx mice had significantly decreased skeletal muscle strength compared to untreated wild type, there were no significant improvements in mdx mice after treatment. Systolic cardiac function, measured as percent shortening fraction, was decreased in untreated mdx mice compared to untreated wild type and there was no significant difference after treatment in mdx mice. Skeletal and cardiac muscle fibrosis were also significantly increased in untreated mdx mice compared to wild type, but there was no significant improvement in treated mdx mice. In exercised dystrophin deficient mice, chronic administration of Tβ4 increased the number of regenerating fibers in skeletal muscle and could have a potential role in treatment of skeletal muscle disease in Duchenne muscular dystrophy.