Molecular Biology

We showed recently that actin assembly by phagosomal membranes facilitates fusion with late endocytic organelles in macrophages. Moreover, lipids that induced phagosomal actin also stimulated this fusion process. In macrophages infected with pathogenic mycobacteria actin-stimulatory lipids led to an increase in pathogen destruction, whereas inhibitors facilitated their growth. A model was proposed whereby phagosomal membrane actin assembly provides tracks for lysosomes to move towards phagosomes, thereby facilitating fusion. Here, we investigated how cAMP affected phagosomal actin assembly in vitro, and phagosomal actin, acidification and late fusion events in J774 macrophages. Latex bead phagosomes are shown to possess adenylyl cyclase activity, which synthesizes cAMP, and phosphodiesterase activity, which degrades cAMP. The system is regulated by protein kinase A (PKA). Increasing cAMP levels inhibited, whereas decreasing cAMP levels stimulated, actin assembly in vitro and within ...

This experiment was undertaken to depict the favourable condition for mycelial growth, molecular identification and phylogenetic relationship of the selected strains of Pleurotus salmoneostramineus. Suitable temperature and pH were obtained at 25ºC and 6, respectively. Mushroom complete, glucose peptone and yeast malt extract culture media were favorable, while Hennerberg and Hoppkins were unfavorable. Dextrin was the best and xylose was the less effective carbon sources. Inorganic nitrogen sources were less effective for the mycelial growth of P. salmoneostramineus. The sequences of internal transcribed spacer (ITS) region of selected strains revealed that the total length ranged from 614 to 663 bp. The size of the ITS1 and ITS2 regions varied among the strains. Sequence analysis showed that 5.8S of rDNA sequences were identical. Phylogenetic tree of the ITS region sequences indicated that strains of P. salmoneostramineus belong to same cluster. The reciprocal homologies of the ITS...

In order to study the capability of a wild-type BTV-2 strain to infect ovine fetuses, 2 groups of 5 pregnant ewes were infected with a wild BTV-2 strain (Group A) or with the same strain passaged once on Culicoides cells (Kc) (Group B). Following infection, EDTA-blood and serum were collected weekly and tested for the presence of BTV RNA/infectious virus and anti-BTV-2 antibodies, respectively. At lambing, precolostral EDTA-blood and serum samples were collected from lambs and tested as before. The lambs were then sampled as scheduled for the dams. All sheep seroconverted by day post-infection 12 and remained seropositive throughout the sampling period (dpi68). BTV was isolated from dpi7 to dpi14 in Group A and from dpi5 to dpi12 in Group B. None of the 14 lambs born had pre-colostral antibodies. Three lambs born of two ewes of Group B were viremic at birth and infectious virus was isolated from blood up to 11 days after birth. By demonstrating that a BTV-2 wt strain passaged only o...

An increase in Ca 2+ influx through L-type Ca 2+ channels is thought to contribute to neuronal dysfunctions that underlie senile symptoms and Alzheimer's disease. The molecular basis of the age-dependent up-regulation in neuronal L-type Ca 2+ channel activity is largely unknown. We show that phosphorylation of the L-type channel Ca v 1.2 by cAMP-dependent protein kinase is increased >2-fold in the hippocampus of aged rats. The hippocampus is critical for learning and is one of the first brain regions to be affected in Alzheimer's disease. Phosphorylation of Ca v 1.2 by cAMP-dependent protein kinase strongly enhances its activity. Therefore, increased Ca v 1.2 phosphorylation may account for a substantial portion of the age-related rise in neuronal Ca 2+ influx and its neuropathological consequences.

Rice stem borer (Scirpophaga sp.) is considered as one of the primary pests causing yield decrease in Indonesia. Genetic engineering is considered as one of the most effective way to improve the resistance of rice plants to yellow stem borer since no resistance gene has been found in rice and its wild relatives. A double T-DNA binary vector p2TDNAcryIAb wich carry two independent T-DNA was used to transform Indonesian elite rice cv Ciherang using Agrobacterium-mediated transformation to generate transgenic rice that free from selectable marker genes. One T-DNA containing a selectable marker hygromycin phosphotransferase (hpt) gene and the other carrying the cry1Ab gene. As many as 37 putative transgenic plants from 14 embryos were generated. PCR analysis demonstrated that 22 out of 37 putative transgenic rice contain cry1Ab gene. All Ciherang rice carrying the cry1Ab gene expressing Cry1Ab protein as shown by the immunostrip assay. Further work will be carried out to determine the t...

Bu caly?mada, dogrudan moment kontrol (DTC) yontemi kullanan bir asenkron motor surucu sistemi analiz edilmi?tir. Bunun icin, asenkron motorun matematiksel d-q modelinden yararlanylarak, DTC kontrollu asenkron motor surucu sisteminin Matlab/Simulink ile benzetimi yapylmy?tyr. DTC yontemi ile kontrol edilen asenkron motorun hem gecici hem de surekli durumda hyzynyn, uretti?i momentin ve stator ile rotor akymlarynyn zamana gore de?i?imleri incelenmi?tir. Surucu sistemin incelenmesinde, motorun belli zaman aralyklarynda bo?ta veya yuk altynda bulundu?u dikkate alynarak hyz, moment ve akym sonuclary elde edilmi?tir. Surucu sistem modelinde bulunan aky ve moment kontrolorlerinin histerezis band geni?likleri evirici anahtarlama frekansyny etkiledi?i ve moment dalgalanmalaryna sebep oldu?u gorulmu?tur. Bu caly?mada, aky ve moment kontrolorlerinin histerezis band geni?likleri aky ve moment dalgalanmalaryna neden oldu?u icin, once uygun band geni?liklerinin secilmesi gerekti?i ve daha sonra ...

Objectives: Human superoxide dismutase 1 (SOD1) is the cytosolic form of this enzyme it detoxifies superoxide anions and attenuates their toxicities and concomitant detrimental effects on the cells. It is believed that the amount of these enzymes present in the oxidative stress-induced diseases is crucial for preventing disease progression. Transfection of rat bone marrow stromal cells (BMSCs) by a constructed vector carrying the human wild-type SOD1 gene, a non-viral gene transfer method, was the main aim of this study. Materials and methods: For this purpose, the rat BMSCs were transfected with the vector using Turbofect reagent and then stabilized. Western-blot and real-time PCR were also used for evaluation of SOD1 expression. Results: Data analysis from RT-PCR and Western-blot techniques revealed that the stable transfected cells could secrete human wild-type SOD1 in the supernatant. Also, the total activity of SOD1 was about 0.5±0.09 U/ml and 0.005±0.002 U/ml in the supernatants of the transfected and not-transfected of rat BMSCs, respectively. Conclusion: This study showed that expansion of the stable transfected rat BMSCs by a constructed vector carrying the human wild-type SOD1 gene is capable of secreting the active SOD1 enzyme under ex-vivo conditions. The recommendation of this study is that the same experiment would be applicable for expression of the other form of this enzyme, SOD3, as well. More valuable information could probably be provided about the variety of the diseases caused by superoxide anions toxicities by intervention and application of the non-viral method for expressions of SOD1 and SOD3 enzymes.

Three human tumor cell lines made resistant to cis-diamminedichloroplatinum(II) (CDDP), SCC-25/CP, MCF-7/CP, and C13, are more sensitive to rhodamine-123 [tetrachloroplatinum(II)] [(PtCl4(Rh-123)2] than are the corresponding parental cell lines. The CDDP-resistant cells have higher intracellular concentrations of PtCl4(Rh-123)2 for the same exposure than do the parent cells. Each of the CDDP-resistant cell lines has an increased level of cytochrome c oxidase activity compared with the parent cell lines, indicating that the resistant cells have greater mitochondrial mass or activity than the parent cells. In fact, there was a linear correlation between the increase in cytochrome c oxidase activity and the increased sensitivity to PtCl4(Rh-123)2 in the CDDP-resistant lines. Exposure of the cells to each of the mitochondrial effectors, chloramphenicol, FCCP, oligomycin, or antimycin prior to and during exposure to CDDP or PtCl4(Rh-123)2 had variable effects on the cytotoxicity of the p...

The establishment of reliable serological methods for cysticercosis in pigs is important for the surveillance, control and prevention of taeniosis/cysticercosis in humans as well as in pigs to prevent economic loss. Both ELISA and immunoblot using glycoproteins (GPs) purified by a single step of preparative iso-electric focusing, which are highly useful for human cysticercosis, have been applied for a serological study in pigs naturally infected withTaenia solium. All sera from pigs showed similar responses to those in human cysticercosis. Therefore, it is expected that both ELISA and immunoblots using GPs would be useful in differentiating infected pigs from uninfected ones.