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Jo?o Miguel Freire Ana Salomé Veiga Thaís M. Concei??o Wioleta Kowalczyk Ronaldo Mohana-Borges David Andreu Nuno C. Santos Andrea T. Da Poian Miguel A. R. B. Castanho 《PloS one》2013,8(12)
Supercharged proteins are a recently identified class of proteins that have the ability to efficiently deliver functional macromolecules into mammalian cells. They were first developed as bioengineering products, but were later found in the human proteome. In this work, we show that this class of proteins with unusually high net positive charge is frequently found among viral structural proteins, more specifically among capsid proteins. In particular, the capsid proteins of viruses from the Flaviviridae family have all a very high net charge to molecular weight ratio (> +1.07/kDa), thus qualifying as supercharged proteins. This ubiquity raises the hypothesis that supercharged viral capsid proteins may have biological roles that arise from an intrinsic ability to penetrate cells. Dengue virus capsid protein was selected for a detailed experimental analysis. We showed that this protein is able to deliver functional nucleic acids into mammalian cells. The same result was obtained with two isolated domains of this protein, one of them being able to translocate lipid bilayers independently of endocytic routes. Nucleic acids such as siRNA and plasmids were delivered fully functional into cells. The results raise the possibility that the ability to penetrate cells is part of the native biological functions of some viral capsid proteins. 相似文献
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The human genome is exposed to oxidative/genotoxic stress by several endogenous and exogenous compounds. These events evoke DNA damage and activate poly(ADP-ribose) polymerase-1 (PARP-1), the key enzyme involved in DNA repair. The massive stress and over-activation of this DNA-bound enzyme can be responsible for an energy crisis and neuronal death. The last data indicated that product of PARP-1, i.e. poly(ADP-ribose) (PAR), acts as a signalling molecule and plays a significant role in nucleus-mitochondria cross-talk. PAR translocated to the mitochondria can be involved in mitochondrial permeability, the release of an apoptosis-inducing factor (AIF). Its translocation into the nucleus leads to chromatin condensation, fragmentation and cell death. The exact mechanism of this novel death pathway has not yet fully been understood. 相似文献
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We determined whether salicylate at pharmacological concentrations inhibits nitric-oxide synthase-2 (NOS-2) and cyclo-oxygenase-2 (COX-2) expressions in RAW 264.7 stimulated with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). Cells were treated with sodium salicylate (10(-7)-10(-4) m) or vehicle for 30 min followed by LPS+IFN-gamma for up to 24 h. Salicylate suppressed NOS-2 and COX-2 protein levels and promoter activities stimulated by LPS+IFN-gamma for 4 h in a concentration-dependent manner but had no effect on NOS-2 expression stimulated by the combined agonists for 24 h. Results from promoter analysis indicate that the binding of CCAAT/enhancer-binding protein beta (C/EBPbeta) to its cognate site at -150/-142 on the NOS-2 promoter region was essential for NOS-2 expression at 4 h but not at 24 h. Salicylate reduced C/EBPbeta binding at 4 h and did not alter its binding at 24 h. NOS-2 and COX-2 protein levels and C/EBPbeta binding stimulated by LPS+IFN-gamma for 4 h were inhibited by a similar battery of signaling inhibitors, suggesting a common pathway for NOS-2 and COX-2 expression. Kinetic analysis indicates that NOS-2, similar to COX-2 expression, at 4 h was largely due to the action of LPS, which induced C/EBPbeta binding, whereas its expression at a longer time point was contributed by IFN-gamma. Our findings implicate two distinct pathways for NOS-2 expression induced by LPS+IFN-gamma. Salicylate at pharmacological concentrations is capable of suppressing the early phase of NOS-2 and COX-2 expression by blocking C/EBPbeta binding. 相似文献
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Marzena Rams-Baron Mateusz Dulski Anna Mrozek-Wilczkiewicz Mateusz Korzec Wioleta Cieslik Ewelina Spaczyńska Piotr Bartczak Alicja Ratuszna Jaroslaw Polanski Robert Musiol 《PloS one》2015,10(6)
New styrylquinoline derivatives with their photophysical constants are described. The synthesis was achieved via Sonogashira coupling using the newly developed heterogeneous nano-Pd/Cu catalyst system, which provides an efficient synthesis of high purity products. The compounds were tested in preliminary fluorescent microscopy studies to in order to identify their preferable cellular localization, which appeared to be in the lipid cellular organelles. The spectroscopic properties of the compounds were measured and theoretical TD-DFT calculations were performed. A biological analysis of the quinolines that were tested consisted of cytotoxicity assays against normal human fibroblasts and colon adenocarcinoma cells. All of the compounds that were studied appeared to be safe and indifferent to cells in a high concentration range. The presented results suggest that the quinoline compounds that were investigated in this study may be valuable structures for development as fluorescent dyes that could have biological applications. 相似文献
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Anna Mrozek-Wilczkiewicz Ewelina Spaczynska Katarzyna Malarz Wioleta Cieslik Marzena Rams-Baron Vladimír Kry?tof Robert Musiol 《PloS one》2015,10(11)
A group of styrylquinolines were synthesized and tested for their anti-proliferative activity. Anti-proliferative activity was evaluated against the human colon carcinoma cell lines that had a normal expression of the p53 protein (HCT116 p53+/+) and mutants with a disabled TP53 gene (HCT116 p53-/-) and against the GM 07492 normal human fibroblast cell line. A SAR study revealed the importance of Cl and OH as substituents in the styryl moiety. Several of the compounds that were tested were found to have a marked anti-proliferative activity that was similar to or better than doxorubicin and were more active against the p53 null than the wild type cells. The cellular localization tests and caspase activity assays suggest a mechanism of action through the mitochondrial pathway of apoptosis in a p53-independent manner. The activity of the styrylquinoline compounds may be associated with their DNA intercalating ability. 相似文献
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Estimation of urinary cotinine cut-off points distinguishing non-smokers, passive and active smokers
Wioleta Zielińska-Danch W?adys?aw Wardas Andrzej Sobczak Izabela Szo?tysek-Bo?dys 《Biomarkers》2007,12(5):484-496
An objective assessment of exposure to tobacco smoke may be accomplished by means of examining particular biomarkers in body fluids. The most common biomarker of tobacco smoke exposure is urinary, or serum, cotinine. In order to distinguish non-smokers from passive smokers and passive smokers from active smokers, it is necessary to estimate cotinine cut-off points. The objective of this article was to apply statistical distribution of urinary cotinine concentration to estimate cut-off points distinguishing the three above-mentioned groups. The examined group consisted of 327 volunteers (187 women and 140 men) who were ethnically homogenous inhabitants of the same urban agglomeration (Sosnowiec, Poland). The values which enabled differentiation of the examined population into groups and subgroups were as follows: 50 µg l-1 (differentiation of non-smokers from passive smokers), 170 µg l-1 (to divide the group of passive smokers into two subgroups: minimally and highly exposed to environmental tobacco smoke), 550 µg l-1 (differentiation of passive smokers from active smokers), and 2100 µg l-1 (to divide group of active smokers into two subgroups: minimally and highly exposed to tobacco smoke). The results suggest that statistical distribution of urinary cotinine concentration is useful for estimating urinary cotinine cut-off points and for assessing the smoking status of persons exposed to tobacco smoke. 相似文献
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Wioleta?Kellmann-Sopy?aEmail author Les?aw?B.?Lahuta Irena?Gie?wanowska Ryszard?J.?Górecki 《Acta Physiologiae Plantarum》2015,37(6):118
The accumulation of soluble carbohydrates in maturing diaspores of flowering plants comprising Arctic populations of Cerastium alpinum, indigenous Antarctic species Colobanthus quitensis and Deschampsia antarctica, and cosmopolitan Poa annua from the Antarctic was investigated. For comparative purposes, the diaspores of two species of flowering plants growing in the area of Olsztyn (Poland), Poa annua (Poaceae) and Cerastium arvense (Caryophyllaceae) were used. A qualitative and quantitative analysis of soluble carbohydrates conducted by means of high-resolution gas chromatography showed that monosaccharides (glucose and fructose), maltose and sucrose, raffinose, myo-inositol and galactinol are ubiquitous in developing and mature diaspores among investigated species. Moreover, D. antarctica and P. annua caryopses additionally contained stachyose and 1-kestose; the seeds of Caryophyllaceae studied were found to contain d-pinitol and d-ononitol. The development and maturation of the seeds of polar Caryophyllaceae and Poaceae were accompanied by the changes in the concentration of their soluble carbohydrates. During maturation, seeds accumulated galactinol and raffinose family of oligosaccharides (RFOs), except C. quitensis. Although seeds of the studied Caryophyllaceae contained d-pinitol and lower amounts of d-ononitol, they did not accumulate α-d-galactoside derivatives of mentioned cyclitols. P. annua caryopses, occurring in the Antarctic, were found to accumulate considerably higher amounts of sucrose and 1-kestose than those developed in Olsztyn. 相似文献
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In C4 plants, such as maize, the photosynthetic apparatus is partitioned over two cell types called mesophyll (M) and bundle sheath (BS), which have different structure and specialization of the photosynthetic thylakoid membranes. We characterized protein phosphorylation in thylakoids of the two cell types from maize grown under either low or high light. Western blotting with phosphothreonine antibodies and ProQ phosphostaining detected light-dependent changes in the protein phosphorylation patterns. LC-MS/MS with alternating CID and electron transfer dissociation sequencing of peptide ions mapped 15 protein phosphorylation sites. Phosphorylated D2, CP29, CP26, Lhcb2 proteins, and ATPsynthase were found only in M membranes. A previously unknown phosphorylation site was mapped in phosphoenolpyruvate carboxykinase from the BS cells. Phosphorylation stoichiometry was calculated from the ratios of normalized ion currents for phosphorylated to nonphosphorylated peptide pairs from the D1, D2, CP43, and PbsH proteins of photosystem II (PSII). Every PSII in M thylakoids contained on average 1.5 ± 0.1 or 2.3 ± 0.2 phosphoryl groups in plants grown under either low or high light, while in BS membranes the corresponding numbers were 0.25 ± 0.1 or 0.7 ± 0.2, respectively. It is suggested that the phosphorylation level, as well as turnover of PSII depend on the structure of thylakoids. 相似文献