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Mutated Version (mutated + version)
Selected AbstractsInterferon regulatory factor-1 acts as a powerful adjuvant in tat DNA based vaccination,JOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2010Arianna Castaldello Genetic vaccines are safe cost-effective approaches to immunization but DNA immunization is an inefficient process. There is, therefore, a pressing need for adjuvants capable of enhancing the immunogenicity and effectiveness of these vaccines. This is particularly important for diseases for which successful vaccines are still lacking, such as cancer and infectious diseases including HIV-1/AIDS. Here we report an approach to enhance the immunogenicity of DNA vaccines involving the use of transcription factors of the Interferon regulatory factor (IRF) family, specifically IRF-1, IRF-3, and IRF-7 using the tat gene as model antigen. Balb/c mice were immunized by three intramuscular inoculations, using a DNA prime-protein boost protocol, with a DNA encoding tat of HIV-1 and the indicated IRFs and immune responses were compared to those induced by vaccination with tat DNA alone. In vivo administration of plasmid DNA encoding IRF-1, or a mutated version of IRF-1 deleted of the DNA-binding domain, enhanced Tat-specific immune responses and shifted them towards a predominant T helper 1-type immune response with increased IFN-, production and cytotoxic T lymphocytes responses. Conversely, the use of IRF-3 or IRF-7 did not affect the tat -induced responses. These findings define IRF-1 and its mutated form as efficacious T helper 1-inducing adjuvants in the context of tat- based vaccination and also providing a new promising candidate for genetic vaccine development. J. Cell. Physiol. 224: 702,709, 2010. © 2010 Wiley-Liss, Inc. [source] Frizzled-1 is involved in the neuroprotective effect of Wnt3a against A, oligomersJOURNAL OF CELLULAR PHYSIOLOGY, Issue 1 2008Marcelo A. Chacón The activation of the canonical Wnt signaling pathway protects hippocampal neurons against the toxicity of Alzheimer's amyloid-,-peptide (A,), however, the role played by the Wnt receptors Frizzleds, has not been studied. We report here that Frizzled-1 mediates the activation of the canonical Wnt/,-catenin pathway by Wnt3a in PC12 cells. In addition, the protective effect of Wnt3a against the toxicity of A, oligomers was modulated by Frizzled-1 expression levels in both PC12 cells and hippocampal neurons. Over-expression of Frizzled-1 significantly increased cell survival induced by Wnt3a and diminished caspase-3 activation, while knocking-down Frizzled-1 expression by antisense oligonucleotides decreased the Wnt3a protection. Over-expression of wild-type ,-catenin, but not a transcriptionally inactive mutated version, prevented the toxicity of A, suggesting that the transcription of Wnt target genes may be involved in these events. This was confirmed by co-transfecting both Frizzled-1 and the inactive form of ,-catenin, which does not elicited protection levels similar to those showed with endogenous ,-catenin. Our results indicate that Wnt3a protects from A,-oligomers toxicity by activating the canonical Wnt signaling pathway through the Frizzled-1 receptor, suggesting a therapeutic potential for this signaling pathway in the treatment of Alzheimer's disease. J. Cell. Physiol. 217: 215,227, 2008. © 2008 Wiley-Liss, Inc. [source] Single amino acid variation in barley 14-3-3 proteins leads to functional isoform specificity in the regulation of nitrate reductaseTHE PLANT JOURNAL, Issue 6 2005Mark P. Sinnige Summary The highly conserved family of 14-3-3 proteins function in the regulation of a wide variety of cellular processes. The presence of multiple 14-3-3 isoforms and the diversity of cellular processes regulated by 14-3-3 suggest functional isoform specificity of 14-3-3 isoforms in the regulation of target proteins. Indeed, several studies observed differences in affinity and functionality of 14-3-3 isoforms. However, the structural variation by which isoform specificity is accomplished remains unclear. Because other reports suggest that specificity is found in differential expression and availability of 14-3-3 isoforms, we used the nitrate reductase (NR) model system to analyse the availability and functionality of the three barley 14-3-3 isoforms. We found that 14-3-3C is unavailable in dark harvested barley leaf extract and 14-3-3A is functionally not capable to efficiently inhibit NR activity, leaving 14-3-3B as the only characterized isoform able to regulate NR in barley. Further, using site directed mutagenesis, we identified a single amino acid variation (Gly versus Ser) in loop 8 of the 14-3-3 proteins that plays an important role in the observed isoform specificity. Mutating the Gly residue of 14-3-3A to the alternative residue, as found in 14-3-3B and 14-3-3C, turned it into a potent inhibitor of NR activity. Using surface plasmon resonance, we show that the ability of 14-3-3A and the mutated version to inhibit NR activity correlates well with their binding affinity for the 14-3-3 binding motif in the NR protein, indicating involvement of this residue in ligand discrimination. These results suggest that both the availability of 14-3-3 isoforms as well as binding affinity determine isoform-specific regulation of NR activity. [source] A dominant nuclear mutation in Chlamydomonas identifies a factor controlling chloroplast mRNA stability by acting on the coding region of the atpA transcriptTHE PLANT JOURNAL, Issue 6 2002Dominique Drapier Summary We have characterized a nuclear mutation, mda1 -ncc1, that affects mRNA stability for the atpA gene cluster in the chloroplast of Chlamydomonas. Unlike all nuclear mutations altering chloroplast gene expression described to date, mda1 -ncc1 is a dominant mutation that still allows accumulation of detectable amounts of atpA mRNAs. At variance with the subset of these mutations that affect mRNA stability through the 5, UTR of a single chloroplast transcript, the mutated version of MDA1 acts on the coding region of the atpA message. We discuss the action of MDA1 in relation to the unusual pattern of expression of atpA that associates particularly short lived-transcripts with a very high translational efficiency. [source] | ||||||||||