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Protein Translation (protein + translation)
Selected AbstractsTranslation of an integral membrane protein in distal dendrites of hippocampal neuronsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 6 2005Jeffrey C. Grigston Abstract Maintenance of synaptic plasticity requires protein translation. Because changes in synaptic strength are regulated at the level of individual synapses, a mechanism is required for newly translated proteins to specifically and persistently modify only a subset of synapses. Evidence suggests this may be accomplished through local translation of proteins at or near synapses in response to plasticity-inducing patterns of activity. A number of proteins important for synaptic function are integral membrane proteins, which require a specialized group of organelles, proteins and enzymatic activities for proper synthesis. Dendrites appear to contain machinery necessary for the proper production of these proteins, and mRNAs for integral membrane proteins have been found localized to dendrites. Experiments are described that investigate the local translation of membrane proteins in the dendrites of cultured rat hippocampal neurons, using fluorescence recovery after photobleaching. Neurons were transfected with cDNAs encoding a fluorescently labeled transmembrane protein, TGN-38. Under conditions where the transport of this reporter construct was inhibited, the appearance of newly synthesized protein was observed via fluorescent microscopy. The dendritic translation of this protein required activation of glutamate receptors. The results demonstrate a functional capacity for activity-dependent synthesis of integral membrane proteins for distal dendrites in hippocampal neurons. [source] Interferon-, synergistically enhances induction of interleukin-6 by double stranded RNA in HeLa cellsFEBS JOURNAL, Issue 9 2000Jennifer L. Harcourt Double stranded RNA (dsRNA), an intermediate that is common during viral infection, directly induces much higher levels of expression of interleukin-6 (IL-6) mRNA than does the cytokine IL-1,. Interferon , (IFN,) by itself does not induce expression of IL-6; nonetheless, IFN, pretreatment dramatically enhances IL-6 induction by dsRNA but not by IL-1,. Mutation of either the activating transcription factor/cyclic AMP response element binding protein (ATF/CREB) or the NF-IL-6 binding element within the IL-6 promoter eliminates most responsiveness of CAT reporter constructs to either dsRNA or to IL-1,. IFN, pretreatment partially restores responsiveness to dsRNA but not to IL-1, when either the ATF/CREB site or the NF-IL-6 site is mutated, but at least one of these sites must be intact for responsiveness to be restored. Mutation of the ,B binding site in the IL-6 promoter eliminates responsiveness to either IL-1, or to dsRNA, and pretreatment with IFN, does not restore any responsiveness. Incubation with dsRNA leads to a decrease in protein translation, especially in cells that have been pretreated with IFN,. Nonetheless, IFN, pretreatment followed by dsRNA leads to very high IL-6 protein levels. These studies demonstrate that major differences exist in the induction of IL-6 at both the mRNA and protein levels by dsRNA compared to cytokines and that IFN, pretreatment selectively enhances IL-6 induction by dsRNA but not by IL-1,. The high levels of IL-6 expression that result when cells encounter class I IFN prior to dsRNA suggest a mechanism for a heightened host response to viral infection with heightened production of this pleotropic cytokine. [source] Cis -preferential recruitment of duck hepatitis B virus core protein to the RNA/polymerase preassembly complexHEPATOLOGY, Issue 1 2002Fritz von Weizsäcker M.D. Hepadnaviral replication requires the concerted action of the polymerase and core proteins to ensure selective packaging of the RNA pregenome into nucleocapsids. Virus assembly is initiated by cis -preferential binding of polymerase to the encapsidation signal ,, present on pregenomic RNA. Using the duck hepatitis B virus (DHBV) model, we analyzed how core protein is recruited to the RNA/polymerase preassembly complex. Two sets of trans-complementation assays were performed in cotransfected hepatoma cells. First, a replication-competent DHBV construct was tested for its ability to rescue replication of genomes bearing mutations within the core region. Self-packaging of wild-type pregenomes was more efficient than cross-packaging of core-deficient pregenomes, and this bias was strongly enhanced if mutant pregenomes coded for self-assembly,competent, but packaging-deficient, core proteins. Second, the site of wild-type core protein translation, i.e., pregenomic RNA (cis) or separate messenger RNA (trans), was analyzed for its effect on the phenotype of a previously described dominant-negative (DN) DHBV core protein mutant. This mutant forms chimeric nucleocapsids with wild-type core proteins and blocks reverse transcription within most, but not all, mixed particles. Strikingly, suppression of viral DNA synthesis by the mutant increased 100-fold when wild-type core protein was provided in trans. Our results suggest that recruitment of core protein to the DHBV preassembly complex occurs in a cis -preferential manner. This mechanism may account for the leakiness of DN DHBV core protein mutants targeting reverse transcription. [source] Hot topics in aging research: protein translation, 2009AGING CELL, Issue 6 2009Brian K. Kennedy Summary In the last few years, links between regulation of mRNA translation and aging have been firmly established in invertebrate model organisms. This year, a possible relationship between mRNA translation and aging in mammals has been established with the report that rapamycin increases lifespan in mice. Other significant findings have connected translation control with other known longevity pathways and provided fodder for mechanistic hypotheses. Here, we summarize advances in this emerging field and raise questions for future studies. [source] Involvement of Gadd153 in the pathogenic action of presenilin-1 mutationsJOURNAL OF NEUROCHEMISTRY, Issue 3 2002Ollivier Milhavet Abstract Mutations in the presenilin-1 (PS1) gene cause early onset familial Alzheimer's disease (FAD) by a mechanism believed to involve perturbed endoplasmic reticulum (ER) function and altered proteolytic processing of the amyloid precursor protein. We investigated the molecular mechanisms underlying cell death and ER dysfunction in cultured cells and knock-in mice expressing FAD PS1 mutations. We report that PS1 mutations cause a marked increase in basal protein levels of the pro-apoptotic transcription factor Gadd153. PS1 mutations increase Gadd153 protein translation without affecting mRNA levels, while decreasing levels of the anti-apoptotic protein Bcl-2. Moreover, an exaggerated Gadd153 response to stress induced by ER stress agents was observed in PS1 mutant cells. Cell death in response to ER stress is enhanced by PS1 mutations, and this endangering effect is attenuated by anti-sense-mediated suppression of Gadd153 production. An abnormality in the translational regulation of Gadd153 may sensitize cells to the detrimental effects of ER stress and contribute to the pathogenic actions of PS1 mutations in FAD. [source] Tissue factor: a mini-reviewJOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE, Issue 3 2007Valéry Daubie Abstract Tissue factor (TF) is historically known as the trigger of the coagulation cascade. This integral membrane glycoprotein forms a ternary complex with factor VIIa (FVIIa) and zymogen factor (FX), which is then activated to factor Xa (FXa). The latter cleaves prothrombin into thrombin (FIIa), which in turn activates fibrinogen in fibrin monomers. What is less known is its additional non-haemostatic roles in inflammation, tumour growth and angiogenesis. This aspect will be developed here. TF, as a transmembrane protein, has a signalling effect requiring FVIIa. TF,FVIIa complex activates G protein-coupled receptor protease-activated receptor 2 (PAR-2) and therefore modulates various cellular processes, such as cell proliferation and survival, gene transcription and protein translation. In this review we will first highlight, using recent structural data, the ,potentially' active domain able to modulate the triggered intracellular response. We also will focus on the still emerging and promising results deciphering the diverse locations in which TF appears. We conclude with a description of an emerging and atypical use of tissue factor in platelet gel surgery for sinus augmentation. Copyright © 2007 John Wiley & Sons, Ltd. [source] Advances in protein turnover analysis at the global level and biological insightsMASS SPECTROMETRY REVIEWS, Issue 5 2010Qingbo Li Abstract The concept of a dynamic state of body constituents, a precursor of the modern term of proteome dynamics, was conceived over a century ago. But, not until recently can we examine the dynamics of individual "constituents" for example, proteins at a truly global level. The path of advancement in our understanding of protein turnover at the global level is marked by the introduction of some key technological innovations. These methods include the isotopic tracer technique in the 1930s, the two-dimensional gel electrophoresis technique in the 1970s, the sector mass spectrometer that could analyze isotopomers of peptides in the early 1990s, the 2D gel/MALDI-TOF proteomics technology in the late 1990s, the booming liquid chromatography/mass spectrometry proteomics technology in this decade, and the recently emerging protein-tagging approaches that offer single-cell resolution for protein turnover measurements. The long-standing inquiry raised in the 1950s about the existence of a dynamic state in different organisms at different physiological conditions can now be answered with an individual "constituent" resolution on a truly global scale. Now it appears that protein degradation is not necessarily an end to the protein function. Rather, it can be the start of a new function because protein degradation clears the way for the action of other proteins. Protein turnover participates in a multi-layer complex regulatory network and shares equal importance with gene transcription and protein translation. The advances in technologies for protein turnover analysis and the improved understanding of the biological role of protein turnover will likely help to solve some long-standing biomedical problems such as the tuberculosis disease that at the present day still affects one-third of the world population. © 2009 Wiley Periodicals, Inc., Mass Spec Rev 29:717,736, 2010 [source] Changes of maternal transcripts in oocytes from persistent follicles in cattleMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2007Brandon M. Lingenfelter Abstract A high incidence of early embryonic loss is associated with prolonged dominance of follicles. The objective of the present experiment was to determine if persistence of a follicle resulted in alterations in mRNA expression of important genes in the oocyte. Cows were assigned to four groups: growing follicles on day 6 (G0h) or day 8 (G48h) and persistent follicles on day 13 (P0h) or day 15 (P48h) of the estrous cycle (estrus,=,day 0). All cows were super-stimulated on day 1,4. Cows in G48h, P0h, and P48h groups received 25 mg prostaglandin (PG) F2, on day 6. Cows in P0h and P48h groups received progesterone from CIDR-B devices on day 5 through 13. Ovaries of cows in G0h, G48h, P0h, and P48h groups were removed on day 6, 8, 13, and 15, respectively. Oocytes were aspirated immediately after colpotomy and denuded of cumulus cells. Quantitative real-time PCR was used to measure the mRNA abundances of 10 selected genes important for early embryogenesis in oocytes obtained from growing and persistent follicles. Relative abundances of MSY2, PARN, and YY1 mRNA (P,<,0.05) were significantly lower in oocytes from persistent than from growing follicles. Oocytes from persistent follicles, however, had greater abundances of PAP and eIF-4E transcripts (P,<,0.05). The data indicate that persistence of a follicle leads to altered abundances of mRNA for genes important for regulation of transcription and protein translation in the oocyte, which could compromise development of early embryos in cows that ovulate a persistent follicle. Mol. Reprod. Dev. © 2006 Wiley-Liss, Inc. [source] Review: The role of microRNAs in kidney diseaseNEPHROLOGY, Issue 6 2010JORDAN YZ LI ABSTRACT MicroRNAs (miRNAs) are short non-coding RNAs that modulate physiological and pathological processes by inhibiting target gene expression via blockade of protein translation or by inducing mRNA degradation. These miRNAs potentially regulate the expression of thousands of proteins. As a result, miRNAs have emerged rapidly as a major new area of biomedical research with relevance to kidney disease. MiRNA expression has been shown to differ between the kidney and other organs as well as between different kidney regions. Furthermore, miRNAs have been found to be functionally important in models of podocyte development, diabetic nephropathy and polycystic kidney disease. Of particular interest, podocyte-specific deletion of Dicer, a key enzyme in the biogenesis of miRNA, results in proteinuria and severe renal impairment in mice. One miRNA (miR-192) can also act as an effector of transforming growth factor-, activity in the high-glucose environment of diabetic nephropathy. Differential expression of miRNAs has been reported in kidney allograft rejection. It is anticipated that future studies involving miRNAs will generate new insights into the complex pathophysiology underlying various kidney diseases, generate diagnostic biomarkers and might be of value as therapeutic targets for progressive kidney diseases. The purpose of this review is to highlight key miRNA developments in kidney diseases and how this might influence the diagnosis and management of patients with kidney disease in the future. [source] Global gene expression profile of Orientia tsutsugamushiPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 8 2010Bon-A Cho Abstract Orientia tsutsugamushi, an obligate intracellular bacterium, is the causative agent of Scrub typhus. The control mechanisms for bacterial gene expression are largely unknown. Here, the global gene expression of O. tsutsugamushi within eukaryotic cells was examined using a microarray and proteomic approaches for the first time. These approaches identified 643 genes, corresponding to approximately 30% of the genes encoded in the genome. The majority of expressed genes belonged to several functional categories including protein translation, protein processing/secretion, and replication/repair. We also searched the conserved sequence blocks (CSBs) in the O. tsutsugamushi genome which is unique in that up to 40% of its genome consists of dispersed repeated sequences. Although extensive shuffling of genomic sequences was observed between two different strains, 204 CSBs, covering 48% of the genome, were identified. When combining the data of CSBs and global gene expression, the CSBs correlates well with the location of expressed genes, suggesting the functional conservation between gene expression and genomic location. Finally, we compared the gene expression of the bacteria-infected fibroblasts and macrophages using microarray analysis. Some major changes were the downregulation of genes involved in translation, protein processing and secretion, which correlated with the reduction in bacterial translation rates and growth within macrophages. [source] Metabolomics: Current technologies and future trendsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 17 2006Katherine Hollywood Abstract The ability to sequence whole genomes has taught us that our knowledge with respect to gene function is rather limited with typically 30,40% of open reading frames having no known function. Thus, within the life sciences there is a need for determination of the biological function of these so-called orphan genes, some of which may be molecular targets for therapeutic intervention. The search for specific mRNA, proteins, or metabolites that can serve as diagnostic markers has also increased, as has the fact that these biomarkers may be useful in following and predicting disease progression or response to therapy. Functional analyses have become increasingly popular. They include investigations at the level of gene expression (transcriptomics), protein translation (proteomics) and more recently the metabolite network (metabolomics). This article provides an overview of metabolomics and discusses its complementary role with transcriptomics and proteomics, and within system biology. It highlights how metabolome analyses are conducted and how the highly complex data that are generated are analysed. Non-invasive footprinting analysis is also discussed as this has many applications to in,vitro cell systems. Finally, for studying biotic or abiotic stresses on animals, plants or microbes, we believe that metabolomics could very easily be applied to large populations, because this approach tends to be of higher throughput and generally lower cost than transcriptomics and proteomics, whilst also providing indications of which area of metabolism may be affected by external perturbation. [source] Retroviral-based gene therapy with cyclooxygenase-2 promotes the union of bony callus tissues and accelerates fracture healing in the ratTHE JOURNAL OF GENE MEDICINE, Issue 3 2008Charles H. Rundle Abstract Background An in vivo gene therapy strategy was developed to accelerate bone fracture repair. Methods Direct injection of a murine leukemia virus-based vector targeted transgene expression to the proliferating periosteal cells arising shortly after fracture. Cyclooxygenase-2 (Cox-2) was selected because the transgene for its prostaglandin products that promote angiogenesis, bone formation and bone resorption, are all required for fracture healing. The human (h) Cox-2 transgene was modified to remove AU-rich elements in the 3,-untranslated region and to improve protein translation. Results In vitro studies revealed robust and sustained Cox-2 protein expression, prostaglandin E2 and alkaline phosphatase production in rat bone marrow stromal cells and osteoblasts transgenic for the hCox-2 gene. In vivo studies in the rat femur fracture revealed that Cox-2 transgene expression produced bony union of the fracture by 21 days post-fracture, a time when cartilage persisted within the fracture tissues of control animals and approximately 1 week earlier than the healing normally observed in this model. None of the ectopic bone formation associated with bone morphogenetic protein gene therapy was observed. Conclusions This study represents the first demonstration that a single local application of a retroviral vector expressing a single osteoinductive transgene consistently accelerated fracture repair. Copyright © 2007 John Wiley & Sons, Ltd. [source] Crystallization and preliminary X-ray diffraction analysis of the middle domain of Paip1ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 10 2009Ahmad Seif Kanaan The poly(A)-binding protein (PABP) simultaneously interacts with the poly(A) tail of mRNAs and the scaffolding protein eIF4G to mediate mRNA circularization, resulting in stimulation of protein translation. PABP is regulated by the PABP-interacting protein Paip1. Paip1 is thought to act as a translational activator in 5, cap-dependent translation by interacting with PABP and the initiation factors eIF4A and eIF3. Here, the crystallization and preliminary diffraction analysis of the middle domain of Paip1 (Paip1M), which produces crystals that diffract to a resolution of 2.2,Å, are presented. [source] Preliminary X-ray crystallographic studies of mouse UPR responsive protein P58(IPK) TPR fragmentACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2008Jiahui Tao Endoplasmic reticulum (ER) stress induces the unfolded protein response (UPR), which can promote protein folding and misfolded protein degradation and attenuate protein translation and protein translocation into the ER. P58(IPK) has been proposed to function as a molecular chaperone to maintain protein-folding homeostasis in the ER under normal and stressed conditions. P58(IPK) contains nine TPR motifs and a C-terminal J-domain within its primary sequence. To investigate the mechanism by which P58(IPK) functions to promote protein folding within the ER, a P58(IPK) TPR fragment without the C-terminal J-domain was crystallized. The crystals diffract to 2.5,Å resolution using a synchrotron X-ray source. The crystals belong to space group P21, with unit-cell parameters a = 83.53, b = 92.75, c = 84.32,Å, , = 90.00, , = 119.36, , = 90.00°. There are two P58(IPK) molecules in the asymmetric unit, which corresponds to a solvent content of approximately 60%. Structure determination by MAD methods is under way. [source] Enhanced IFN, production in adenosine-treated CHOCells: A mechanistic studyBIOTECHNOLOGY PROGRESS, Issue 3 2009William P. K. Chong Abstract Adenosine causes growth arrest in recombinant mammalian cell cultures, which results in enhanced productivity of the recombinant protein. Adenosine is also known to increase intracellular ATP level when added to mammalian cells. As a cell's energy level affects its protein expression capacity, we investigated the factors that contribute to the increase in recombinant protein productivity. Chinese hamster ovary (CHO) cells expressing human interferon-gamma (IFN,) were treated with 1 mM adenosine on Day 2 of culture. The growth arrest resulted in 60% reduction in integral viable cell density when compared with control. However, IFN, titer improved 1.4-fold alongside a 2.5-fold increase in average specific productivity. The adenosine-treated cells also experienced a two-fold increase in ATP level that sustained for 3 days. Western blot studies revealed a relatively short-lived but strong activation of the energy sensor AMP-activated protein kinase (AMPK) in adenosine-treated cells. Activation of AMPK was probably due to adenosine being temporarily converted to AMP. Activated AMPK should have down-regulated protein translation by preventing mammalian target of rapamycin (mTOR) from phosphorylating and inactivating 4E-binding protein 1 (4E-BP1), a key repressor of protein translation initiation. However, Western blots showed increased phosphorylation of 4E-BP1 on Day 2 that lasted 3 days. This implied that a high concentration of ATP could keep 4E-BP1 inhibited, probably by directly modulating mTOR. This corroborated with an earlier in vitro observation (Dennis et al., Science. 2001;294:1102-1105). Inhibition of translation initiation repression is thus likely to contribute in part to the improvement in IFN,-specific productivity and titer. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source] Hyperosmotic Stress in Murine Hybridoma Cells: Effects on Antibody Transcription, Translation, Posttranslational Processing, and the Cell CycleBIOTECHNOLOGY PROGRESS, Issue 2 2004Zhe Sun Mechanisms for increased antibody production in batch cultures of murine hybridoma cells in response to hyperosmotic stress were investigated. The rates of immunoglobulin transcription and protein translation and posttranslational processing were determined in control and hyperosmotic cultures. Changes in immunoglobulin transcription played a minor role in the increase in antibody production in response to hyperosmotic stress. In contrast, protein translation increased substantially in response to osmotic stress. However, the antibody translation rate remained relatively constant after correcting for the overall increase in protein translation. Cell size and intracellular antibody pool also increased in response to hyperosmolarity. The intracellular antibody pool increased proportionately with the increase in cell size, indicating that hyperosmotic cultures do not selectively increase their intracellular antibody population. Changes in cell cycle distribution in response to osmotic stress and the relationship between the cell cycle and antibody production were also evaluated. Hyperosmotic stress altered the cell cycle distribution, increasing the fraction of the cells in S-phase. However, this change was uncorrelated with the increase in antibody production rate. Immunoglobulin degradation was relatively low (,15%) and remained largely unchanged in response to hyperosmotic stress. There was no apparent increase in immunoglobulin stability as a result of osmotic stress. Antibody secretion rates increased approximately 50% in response to osmotic stress, with a commensurate increase in the antibody assembly rate. The rate of transit through the entire posttranslational processing apparatus increased, particularly for immunoglobulin light chains. The levels of endoplasmic reticulum chaperones did not increase as a fraction of the total cellular protein but were increased on a per cell basis as the result of an increase in total cellular protein. A difference in the interactions between the immunoglobulin heavy chains and BiP/GRP78 was observed in response to hyperosmotic conditions. This change in interaction may be correlated with the decrease in transit time through the posttranslational pathways. The increase in the posttranslational processing rate appears to be commensurate with the increase in antibody production in response to hyperosmotic stress. 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