Ku heterodimer binds to both ends of the Werner protein and functional interaction occurs at the Werner N-terminus

The human Werner syndrome protein, WRN, is a member of the RecQ helicase family and contains 3′→5′ helicase and 3′→5′ exonuclease activities. Recently, we showed that the exonuclease activity of WRN is greatly stimulated by the human Ku heterodimer protein. We have now mapped this interaction physically and functionally. The Ku70 subunit specifically interacts with the N-terminus (amino acids 1–368) of WRN, while the Ku80 subunit interacts with its C-terminus (amino acids 940– 1432). Binding between Ku70 and the N-terminus of WRN (amino acids 1–368) is sufficient for stimulation of WRN exonuclease activity. A mutant Ku heterodimer of full-length Ku80 and truncated Ku70 (amino acids 430–542) interacts with C-WRN but not with N-WRN and cannot stimulate WRN exonuclease activity. This emphasizes the functional significance of the interaction between the N-terminus of WRN and Ku70. The interaction between Ku80 and the C-terminus of WRN may modulate some other, as yet unknown, function. The strong interaction between Ku and WRN suggests that these two proteins function together in one or more pathways of DNA metabolism.     

Contributions of Ca2+ to galectin-1-induced exposure of phosphatidylserine on activated neutrophils

Apoptotic cells redistribute phosphatidylserine (PS) to the cell surface by both Ca(2+)-dependent and -independent mechanisms. Binding of dimeric galectin-1 (dGal-1) to glycoconjugates on N-formyl-Met-Leu-Phe (fMLP)-activated neutrophils exposes PS and facilitates neutrophil phagocytosis by macrophages, yet it does not initiate apoptosis. We asked whether dGal-1 initiated Ca(2+) fluxes that are required to redistribute PS to the surface of activated neutrophils. Prolonged occupancy by dGal-1 was required to maximally mobilize PS to the surfaces of fMLP-activated neutrophils. Like fMLP, dGal-1 rapidly elevated cytosolic Ca(2+) levels in Fluo-4-loaded neutrophils. An initial Ca(2+) mobilization from intracellular stores was followed by movement of extracellular Ca(2+) to the cytosolic compartment, with return to basal Ca(2+) levels within 10 min. Chelation of extracellular Ca(2+) did not prevent PS mobilization. Chelation of intracellular Ca(2+) revealed that fMLP and dGal-1 independently release Ca(2+) from intracellular stores that cooperate to induce optimal redistribution of PS. Ca(2+) mobilization by ionomycin did not permit dGal-1 to mobilize PS, indicating that fMLP initiated both Ca(2+)-dependent and -independent signals that facilitated dGal-1-induced exposure of PS. dGal-1 elevated cytosolic Ca(2+) and mobilized PS through a pathway that required action of Src kinases and phospholipase Cgamma. These results demonstrate that transient Ca(2+) fluxes contribute to a sustained redistribution of PS on neutrophils activated with fMLP and dGal-1.     

Evaluation of tomato genotypes for resistance to whitefly (Bemisia tabaci Gennadius) and tomato leaf curl virus in eastern India

Tomato production in the tropics is threatened by whitefly infestation and tomato leaf curl virus (ToLCV) causing severe economic losses. No stable resistance to these biotic challenges has been identified in eastern India. Therefore, initial screening of 19 advance breeding lines of tomato was carried out during the year 2016–17. Based on the whitefly population per leaf and tomato leaf curl disease severity, eight tomato genotypes were selected for final screening during the year 2017–18. Morphological leaf traits and biochemical parameters in tomato leaf were assessed in selected genotypes and considered as potential mediators of resistance. Significant variation was observed for whitefly infestation and ToLCV disease severity among the tomato genotypes tested. Higher leaf trichome density with narrow and thinner leaves and higher total phenol content in leaf emerged as reliable morphological and biochemical markers for early selection of tomato genotype resistant to whitefly and ToLCV disease. The resistance of tomato genotypes 2016/Res-1, 2015/Res-5, 2014/Res-1 and 2014/Res-4 to both whitefly and ToLCV disease is based on antixenotic properties and they could be utilized in future breeding to enhance stable resistance against these biotic challenges.     

A comparative study of biological potentiality and EAC cell growth inhibition activity of Phyllanthus acidus (L.) fruit pulp and seed in Bangladesh

Medicinal plant-derived bioactive compounds have recently gained more interest in biological research as an important source of novel drug candidates. Phyllanthus acidus (L.) is a widely distributed herbal medicinal plant naturally used in Ayurvedic medicine in Bangladesh. The present study focused on exploring the biological potential as well as the inhibitory effect of EAC cell growth with a comparative analysis between Phyllanthus acidus fruit pulp and seed. Crude methanol extract of P. acidus (MEPA) fruit pulp and seed was assessed as DPPH and NO free radical scavengers. While Brine Shrimp lethality bioassay, the standard protocol of phytochemical screening and hemagglutination assay were performed successively to determine the toxic effect on normal cells, the identification of some crucial phytochemicals, and the existence of lectin protein. EAC (Ehrlich’s Ascites Carcinoma) cell growth inhibition was determined by hemocytometer and morphological changes of EAC cells were observed by a fluorescence microscope using Swiss albino mice. The IC₅₀ value of MEPA fruit pulp and seed was obtained as 57.159 µg/ml and 288.743 µg/ml respectively where minimal toxic effects on Brine Shrimp nauplii demonstrates that it is a good source of natural antioxidant compounds. Again, MEPA fruit pulp and seed-mediated effective agglutination of mouse blood erythrocyte strongly support the presence of lectin protein. Furthermore, MEPA fruit pulp and seed extract-treated EAC cells showed 65.71% and 28.57% growth inhibition respectively. The fluorescent microscopic examination of EAC cells treated with MEPA fruit pulp has shown more remarkable structural changes in the nucleus than that of seed. Based on the above findings, the present study reveals that MEPA fruit pulp can be considered as a novel biological candidate for the treatment of fatal diseases shortly.     

Orientation enhancement in early visual processing can explain time course of brightness contrast and White’s illusion

Dynamics of orientation tuning in V1 indicates that computational model of V1 should not only comprise of bank of static spatially oriented filters but also include the contribution for dynamical response facilitation or suppression along orientation. Time evolution of orientation response in V1 can emerge due to time- dependent excitation and lateral inhibition in the orientation domain. Lateral inhibition in the orientation domain suggests that Ernst Mach’s proposition can be applied for the enhancement of initial orientation distribution that is generated due to interaction of visual stimulus with spatially oriented filters and subcortical temporal filter. Oriented spatial filtering that appears much early ( $<$ 70 ms) in the sequence of visual information processing can account for many of the brightness illusions observed at steady state. It is therefore expected that time evolution of orientation response might be reflecting in the brightness percept over time. Our numerical study suggests that only spatio-temporal filtering at early phase can explain experimentally observed temporal dynamics of brightness contrast illusion. But, enhancement of orientation response at early phase of visual processing is the key mechanism that can guide visual system to predict the brightness by “Max-rule” or “Winner Takes All” (WTA) estimation and thus producing White’s illusions at any exposure.     

The protective role of metformin in autophagic status in peripheral blood mononuclear cells of type 2 diabetic patients

Autophagy plays an important role in the pathophysiology of type 2 diabetes (T2D). Metformin is the most common antidiabetic drug. The main objective of this study was to explore the molecular mechanism of metformin in starvation‐induced autophagy in peripheral blood mononuclear cells (PBMCs) of type 2 diabetic patients. PBMCs were isolated from 10 diabetic patients and 7 non‐diabetic healthy volunteers. The autophagic puncta and markers were measured with the help of monodansylcadaverine staining and western blot. Additionally, transmission electron microscopy was also performed. No significant changes were observed in the initial autophagy marker protein levels in PBMCs of T2D after metformin treatment though diabetic PBMCs showed a high level of phospho‐mammalian target of rapamycin, p62 and reduced expression of phospho‐AMP‐activated protein kinase and lysosomal membrane‐associated protein 2, indicating a defect in autophagy. Also, induction of autophagy by tunicamycin resulted in apoptosis in diabetic PBMCs as observed by caspase‐3 cleavage and reduced expression of Bcl2. Inhibition of autophagy by bafilomycin rendered consistent expression of p62 indicating a defect in the final process of autophagy. Further, electron microscopic studies also confirmed massive vacuole overload and a sign of apoptotic cell death in PBMCs of diabetic patients, whereas metformin treatment reduced the number of autophagic vacuoles perhaps by lysosomal fusion. Thus, our results indicate that defective autophagy in T2D is associated with the fusion process of lysosomes which could be overcome by metformin.     

SU5416 and EGCG Work Synergistically and Inhibit Angiogenic and Survival Factors and Induce Cell Cycle Arrest to Promote Apoptosis in Human Malignant Neuroblastoma SH-SY5Y and SK-N-BE2 Cells

Malignant neuroblastomas are solid tumors in children. Available therapeutic agents are not highly effective for treatment of malignant neuroblastomas. Therefore, new treatment strategies are urgently needed. We tested the efficacy of combination of SU5416 (SU), an inhibitor of the vascular endothelial growth factor receptor-2 (VEGFR-2), and (−)-epigallocatechin-3-gallate (EGCG), a polyphenolic compound from green tea, for controlling growth of human malignant neuroblastoma SH-SY5Y and SK-N-BE2 cells. Combination of 20 μM SU and 50 μM EGCG synergistically inhibited cell survival, suppressed expression of VEGFR-2, inhibited cell migration, caused cell cycle arrest, and induced apoptosis. Combination of SU and EGCG effectively blocked angiogenic and survival pathways and modulated expression of cell cycle regulators. Apoptosis was induced by down regulation of Bcl-2, activation of caspase-3, and cleavage of the DNA repair enzyme poly(ADP-ribose) polymerase (PARP). Taken together, this combination of drugs can be a promising therapeutic strategy for controlling the growth of human malignant neuroblastoma cells.