Inhibition of oxidative stress-induced amyloid β formation in NT2 neurons by culture filtrate of a strain of Streptomyces antibioticus

Actinomycetes isolated from Iran soil habitats were tested for the capacity to produce compounds which can protect neurons from cell death generated by oxidative stress in NT2 neurons. Confirmation of our initial hit was accomplished via the determination of amyloid β level using the enzyme-linked immunosorbent assay test. The most interesting amyloid β formation inhibitor discovered in our study was a secondary metabolite which was produced by strain HM45. This bioactive strain was identified as a strain of Streptomyces antibioticus DSM 40234 using polyphasic approach. The strain HM45 was deposited in Deutsche Sammlung von Mikroorganismen und Zellkulturen as S. antibioticus DSM 41955 and University of Tehran Microorganisms Sollection as S. antibioticus UTMC 00105. This work is the first report on efficiency of an actinomycete metabolite in prohibition of neurons death caused by amyloid β formation.     

Physiological responses to NaCl stress in three wild species of potato in vitro

In this study, responses of wild species of potato to NaCl stress were investigated in vitro. In S. stoloniferum and S. bulbosum, length of the shoot, fresh and dry weight, photosynthetic pigments, K⁺ concentration, K⁺/Na⁺ ratio, ascorbate pool, anthocyanin, and phenolic and flavonoid compounds were decreased in response to salinity. In these species, salinity increased the level of Na⁺, lipid peroxidation, proline and ion leakage percentage. In S. acaule, the length of the shoot, and fresh and dry weight were not affected by salinity. Photosynthetic pigments, Na⁺ concentration, proline, flavonoid and phenolic compounds quantities were increased and K⁺/Na⁺ ratio were decreased. K⁺ concentration, lipid peroxidation, ascorbate pool, anthocyanin and ion leakage were not changed by NaCl stress. Superoxide dismutase, guaiacol peroxidase, ascorbate peroxidase and catalase activities were increased in all species. The results suggest that the non-enzymatic antioxidant capacity in S. acaule (salt tolerant) is more important than the enzymatic antioxidant capacity in comparison with the other species.     

Increased expression of recombinant human tissue plasminogen activator in Leishmania tarentolae

Recombinant tissue plasminogen activator (rt-PA) is one of the most important thrombolytic agents for treating cardiovascular obstructions such as stroke. Glycoprotein rt-PA is a serine protease, consisting of 527 amino acids of which 35 are cysteine residues. A variety of recombinant protein expression systems have been developed for heterologous gene expression in prokaryotic and eukaryotic hosts. In recent years, Leishmania tarentolae has been considered because of its safety aspects and special attributes in expression of complex proteins. In this study, two expression cassettes, each one including two copies of t-PA cDNA, were used for integration into the L. tarentolae genome by electroporation. Transformed clones were selected in the presence of appropriate antibiotics. Expression of active rt-PA was confirmed by Western blot and Zymography tests. Real-time PCR analysis was applied to investigate the presence of multiple t-PA gene copies in the parasite genome. Correlation of t-PA gene dosage and production rate was confirmed with real-time PCR. It was shown that the expression level of rt-PA in L. tarentolae is at least 480 IU/mL of culture media. This concentration of rt-PA is seven times higher than what was reported in previous studies in L. tarentolae and some other eukaryotic systems.     

Long-term stability of marker gene expression in Prunus subhirtella: a model fruit tree species

Transgenic trees currently are being produced by Agrobacterium-mediated transformation and biolistics. Since trees are particularly suited for long-term evaluations of the impact of the technology, Prunus subhirtella autumno rosa (PAR) was chosen as model fruit tree species and transformed with a reporter gene (uidA) under the control of the 35S promoter. Using Southern and GUS fluorometric techniques, we compared transgene copy numbers and observed stability of transgene expression levels in 34 different transgenic plants, grown under in vitro, greenhouse and screenhouse conditions, over a period of 9 years. An influence of grafting on gene expression was not observed. No silenced transgenic plant was detected. Overall, these results suggest that transgene expression in perennial species, such as fruit trees, remains stable in time and space, over extended periods and in different organs, confirming the value of PAR as model species to study season-dependent regulation in mature stone fruit tissues. While the Agrobacterium-derived Prunus transformants contained one to two copies of the transgenes, 91% of the transgenic events also contained various lengths of the bacterial plasmid backbone, indicating that the Agrobacterium-mediated transformation is not as precise as previously perceived. The implications for public acceptance and future applications are discussed.     

Distinct functions of the Drosophila Nup153 and Nup214 FG domains in nuclear protein transport

The phenylanine-glycine (FG)-rich regions of several nucleoporins both bind to nuclear transport receptors and collectively provide a diffusion barrier to the nuclear pores. However, the in vivo roles of FG nucleoporins in transport remain unclear. We have inactivated 30 putative nucleoporins in cultured Drosophila melanogaster S2 cells by RNA interference and analyzed the phenotypes on importin alpha/beta-mediated import and CRM1-dependent protein export. The fly homologues of FG nucleoporins Nup358, Nup153, and Nup54 are selectively required for import. The FG repeats of Nup153 are necessary for its function in transport, whereas the remainder of the protein maintains pore integrity. Inactivation of the CRM1 cofactor RanBP3 decreased the nuclear accumulation of CRM1 and protein export. We report a surprisingly antagonistic relationship between RanBP3 and the Nup214 FG region in determining CRM1 localization and its function in protein export. Our data suggest that peripheral metazoan FG nucleoporins have distinct functions in nuclear protein transport events.     

Simultaneous purification and polymerization method for bovine serum albumin preparation

Bovine serum albumin (BSA) has various applications in blood group serology and different research purposes. In this study purification of BSA has been compared with human serum albumin (HSA) using modified ethanol precipitation method based on the method of Cohn. The purification process was carried out under controlled conditions, particularly of ethanol concentration, pH, ionic strength and temperature. It was revealed that the produced BSA and HSA have purity more than 95%. It is obvious that HSA can be used, as a drug when the amount of its polymers is less than 5% whereas polymer generation is required in order to enhance the potentiating properties of BSA in agglutination of red cells. We propose here a simple and rapid two-step method for simultaneously purification and polymerization of BSA. By this method simply BSA with desired amount of polymers was obtained by 40% ethanol concentration.     

Salicylic acid inducible nucleocytoplasmic shuttling of NPR1 fusion proteins in human cells

Ligand inducible proteins that enable precise and reversible control of nuclear translocation of passenger proteins have broad applications ranging from genetic studies in mammals to therapeutics that target diseases such as cancer and diabetes. One of the drawbacks of the current translocation systems is that the ligands used to control nuclear localization are either toxic or prone to crosstalk with endogenous protein cascades within live animals. We sought to take advantage of salicylic acid (SA), a small molecule that has been extensively used in humans. In plants, SA functions as a hormone that can mediate immunity and is sensed by the nonexpressor of pathogenesis-related (NPR) proteins. Although it is well recognized that nuclear translocation of NPR1 is essential to promoting immunity in plants, the exact subdomain of Arabidopsis thaliana NPR1 (AtNPR1) essential for SA-mediated nuclear translocation is controversial. Here, we utilized the fluorescent protein mCherry as the reporter to investigate the ability of SA to induce nuclear translocation of the full-length NPR1 protein or its C-terminal transactivation (TAD) domain using HEK293 cells as a heterologous system. HEK293 cells lack accessory plant proteins including NPR3/NPR4 and are thus ideally suited for studying the impact of SA-induced changes in NPR1. Our results obtained using a stable expression system show that the TAD of AtNPR1 is sufficient to enable the reversible SA-mediated nuclear translocation of mCherry. Our studies advance a basic understanding of nuclear translocation mediated by the TAD of AtNPR1 and uncover a biotechnological tool for SA-mediated nuclear localization.     

Recognition of functional genetic polymorphism using ESE motif definition: a conservative evolutionary approach to CYP2D6/CYP2C19 gene variants

Genetica - Although predicting the effects of variants near intron-exon boundaries is relatively straightforward, predicting the functional Exon Splicing Enhancers (ESEs) and the possible effects...     

2q35-rs13387042 variant and the risk of breast cancer: a case–control study

Molecular Biology Reports - Breast Cancer is the most frequent neoplasm diagnosed among women worldwide. Genetic background and lifestyle/environment play a significant role in the disease...