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RNAi Technology (rnai + technology)
Selected AbstractsOverexpression and significance of prion protein in gastric cancer and multidrug-resistant gastric carcinoma cell line SGC7901/ADRINTERNATIONAL JOURNAL OF CANCER, Issue 2 2005Jingping Du Abstract In our previous work, cellular prion protein (PrPc) was identified as an upregulated gene in adriamycin-resistant gastric carcinoma cell line SGC7901/ADR compared to its parental cell line SGC7901. Here we investigate the expression of PrPc in gastric cancer and whether it was involved in multidrug resistance (MDR) of gastric cancer. We demonstrated that PrPc was ubiquitously expressed in gastric cancer cell lines and tissues. PrPc conferred resistance of both P-glycoprotein (P-gp)-related and P-gp-nonrelated drugs on SGC7901, which was accompanied by decreased accumulation and increased releasing amount of adriamycin in PrPc-overexpressing cell line. Inhibition of PrPc expression by antisense or RNAi technology could partially reverse multidrug-resistant phenotype of SGC7901/ADR. PrPc significantly upregulated the expression of the classical MDR-related molecule P-gp but not multidrug resistance associated protein and glutathione S-transferase pi. The PrPc-induced MDR could be partially reversed by P-gp inhibitor verapamil. PrPc could also suppress adriamycin-induced apoptosis and alter the expression of Bcl-2 and Bax, which might be another pathway contributing to PrPc-related MDR. The further study of the biological functions of PrPc may be helpful for understanding the mechanisms of occurrence and development of clinical gastric carcinoma and PrPc-related MDR and developing possible strategies to treat gastric cancer. [source] Active caspase-3 is required for osteoclast differentiationJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2006K.H. Szymczyk Based on our earlier observation that caspase-3 is present in osteoclasts that are not undergoing apoptosis, we investigated the role of this protein in the differentiation of primary osteoclasts and RAW264.7 cells (Szymczyk KH, et al., 2005, Caspase-3 activity is necessary for RANKL-induced osteoclast differentiation. The Proceedings of the 8th ICCBMT). We noted that osteoclast numbers are decreased in long bones of procaspase-3 knockout mice and that receptor activator of NF-,B ligand (RANKL) does not promote differentiation of isolated preosteoclasts. In addition, after treatment with inhibitors of caspase-3 activity, neither the wild-type primary nor the RAW264.7 cells express TRAP or became multinucleated. We found that immediately following RANKL treatment, procaspase-3 is cleaved and the activated protein is localized to lipid regions of the plasma membrane and the cytosol. We developed RAW264.7 procaspase-3 knockdown clonal cell lines using RNAi technology. Again, treatment with RANKL fails to induce TRAP activity or multinucleation. Finally, we evaluated NF-,B in procaspase-3 silenced cells. We found that RANKL treatment prevented activation and nuclear translocation of NF-,B. Together these findings provide direct support for the hypothesis that caspase-3 activity is required for osteoclast differentiation. J. Cell. Physiol. 209: 836,844, 2006. © 2006 Wiley-Liss, Inc. [source] Recent progress in the development of RNA interference for plant parasitic nematodesMOLECULAR PLANT PATHOLOGY, Issue 5 2007CATHERINE J. LILLEY SUMMARY RNA interference (RNAi), first described for Caenorhabditis elegans, has emerged as a powerful gene silencing tool for investigating gene function in a range of organisms. Recent studies have described its application to plant parasitic nematodes. Genes expressed in a range of cell types are silenced when preparasitic juvenile nematodes take up double-stranded (ds)RNA that elicits a systemic RNAi response. Important developments over the last year have shown that in planta expression of a dsRNA targeting a nematode gene can successfully induce silencing in parasitizing nematodes. When the targeted gene has an essential function, a resistance effect is observed paving the way for the potential use of RNAi technology to control plant parasitic nematodes. [source] Alleviating peanut allergy using genetic engineering: the silencing of the immunodominant allergen Ara h 2 leads to its significant reduction and a decrease in peanut allergenicityPLANT BIOTECHNOLOGY JOURNAL, Issue 2 2008Hortense W. Dodo Summary Peanut allergy is one of the most life-threatening food allergies and one of the serious challenges facing the peanut and food industries. Current proposed solutions focus primarily on ways to alter the immune system of patients allergic to peanut. However, with the advent of genetic engineering novel strategies can be proposed to solve the problem of peanut allergy from the source. The objectives of this study were to eliminate the immunodominant Ara h 2 protein from transgenic peanut using RNA interference (RNAi), and to evaluate the allergenicity of resulting transgenic peanut seeds. A 265-bp-long PCR product was generated from the coding region of Ara h 2 genomic DNA, and cloned as inverted repeats in pHANNIBAL, an RNAi-inducing plant transformation vector. The Ara h 2-specific RNAi transformation cassette was subcloned into a binary pART27 vector to construct plasmid pDK28. Transgenic peanuts were produced by infecting peanut hypocotyl explants with Agrobacterium tumefaciens EHA 105 harbouring the pDK28 construct. A total of 59 kanamycin-resistant peanut plants were regenerated with phenotype and growth rates comparable to wild type. PCR and Southern analyses revealed that 44% of plants stably integrated the transgene. Sandwich ELISA performed using Ara h 2-mAbs revealed a significant (P < 0.05) reduction in Ara h 2 content in several transgenic seeds. Western immunobloting performed with Ara h 2-mAb corroborated the results obtained with ELISA and showed absence of the Ara h 2 protein from crude extracts of several transgenic seeds of the T0 plants. The allergenicity of transgenic peanut seeds expressed as IgE binding capacity was evaluated by ELISA using sera of patients allergic to peanut. The data showed a significant decrease in the IgE binding capacity of selected transgenic seeds compared to wild type, hence, demonstrating the feasibility of alleviating peanut allergy using the RNAi technology. [source] Engineering photosynthetic light capture: impacts on improved solar energy to biomass conversionPLANT BIOTECHNOLOGY JOURNAL, Issue 6 2007Jan H. Mussgnug Summary The main function of the photosynthetic process is to capture solar energy and to store it in the form of chemical ,fuels'. Increasingly, the photosynthetic machinery is being used for the production of biofuels such as bio-ethanol, biodiesel and bio-H2. Fuel production efficiency is directly dependent on the solar photon capture and conversion efficiency of the system. Green algae (e.g. Chlamydomonas reinhardtii) have evolved genetic strategies to assemble large light-harvesting antenna complexes (LHC) to maximize light capture under low-light conditions, with the downside that under high solar irradiance, most of the absorbed photons are wasted as fluorescence and heat to protect against photodamage. This limits the production process efficiency of mass culture. We applied RNAi technology to down-regulate the entire LHC gene family simultaneously to reduce energy losses by fluorescence and heat. The mutant Stm3LR3 had significantly reduced levels of LHCI and LHCII mRNAs and proteins while chlorophyll and pigment synthesis was functional. The grana were markedly less tightly stacked, consistent with the role of LHCII. Stm3LR3 also exhibited reduced levels of fluorescence, a higher photosynthetic quantum yield and a reduced sensitivity to photoinhibition, resulting in an increased efficiency of cell cultivation under elevated light conditions. Collectively, these properties offer three advantages in terms of algal bioreactor efficiency under natural high-light levels: (i) reduced fluorescence and LHC-dependent heat losses and thus increased photosynthetic efficiencies under high-light conditions; (ii) improved light penetration properties; and (iii) potentially reduced risk of oxidative photodamage of PSII. [source] |