Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A manifestation of Alzheimer's disease is the presence of amyloid depositions in brains of afflicted individuals. A major component of these depositions is the amyloid beta-protein, which is a truncated form of the larger amyloid beta-protein precursor (APP). To investigate the regulation of APP gene expression, the APP promoter and selected deletions were placed 5' to the reporter gene chloramphenicol acetyltransferase. The promoter deletions were transfected into different cell lines that showed variant levels of endogenous APP transcripts. Transient transfection assays showed that 96 base pairs 5' to the transcriptional start site are sufficient for cell type-specific promoter activity. A nuclear factor that binds to this region in a sequence-specific manner was identified by mobility shift electrophoresis, DNase footprinting, and methylation interference. The DNase-protected region covers about 25 base pairs on both strands (position -31 to -55). Mutations within this domain revealed a sequence of 12 base pairs that is crucial for factor binding. This sequence overlaps with the consensus sequences for transcription factors AP-1 and AP-4. However, competition experiments suggest that the nuclear factor that binds to the APP promoter is distinct from both AP-1 and AP-4. Factor binding to the characterized recognition sequence is observed in nuclear extracts originating from human, mouse, and rat cells, suggesting a high degree of conservation.
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PMID:The amyloid beta-protein precursor promoter. A region essential for transcriptional activity contains a nuclear factor binding domain. 138 Sep 60

We cloned, sequenced and characterized a promoter region of the mouse homologue of the Alzheimer's disease amyloid precursor protein (APP)-encoding gene. The promoter region is highly homologous to that of the human APP (hAPP) gene. It has a high G+C content, lacks typical 'TATA' and 'CAAT' boxes, and contains possible binding sites for AP-1, heat-shock factor, Sp1 and AP-4. The promoter region was fused with the cat reporter gene, and the fusion genes were transfected to both the NB41A3 (mouse neuroblastoma) and L-cell lines. The promoter activity was monitored by chloramphenicol acetyltransferase (CAT) activity in a transient expression assay. The promoter was equally active in both cell lines. The deletion analysis revealed that there existed a negative regulatory element(s) between -153 and -100 bp and a positive element(s) between -100 and -37 bp. The negative element was shown to suppress the transcriptional activity of heterologous simian virus 40 promoter. DNase I footprinting experiments revealed that three nuclear protein-binding sites existed in the regulatory region, one in the negative and two in the positive regulatory regions. Gel retardation assay showed that Sp1 was one of the factors binding to the positive regulatory region. A nuclear factor binding to the negative regulatory region seemed to be missing in brain.
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PMID:Positive and negative regulatory elements for the expression of the Alzheimer's disease amyloid precursor-encoding gene in mouse. 155 68

Transgenic mice carrying heterologous genes directed by a 670-bp segment of the regulatory sequence from the human transferrin (TF) gene demonstrated high expression in brain. Mice carrying the chimeric 0.67kbTF-CAT gene expressed TF-CAT in neurons and glial cells of the nucleus basalis, the cerebrum, corpus callosum, cerebellum, and hippocampus. In brains from two independent TF-CAT transgenic founder lines, copy number of TF-CAT mRNA exceeded the number of mRNA transcripts encoding either mouse endogenous transferrin or mouse endogenous amyloid precursor protein. In two transgenic founder lines, the chloramphenicol acetyltransferase (CAT) protein synthesized from the TF-CAT mRNA was estimated to be 0.10-0.15% of the total soluble proteins of the brain. High expression observed in brain indicates that the 0.67kbTF promoter is a promising director of brain expression of heterologous genes. Therefore, the promoter has been used to express the three common human apolipoprotein E (apoE) alleles in transgenic mouse brains. The apoE alleles have been implicated in the expression of Alzheimer disease, and the human apoE isoforms are reported to interact with different affinities to the brain beta-amyloid and tau protein in vitro. Results of this study demonstrate high expression and production of human apoE proteins in transgenic mouse brains. The model may be used to characterize the interaction of human apoE isoforms with other brain proteins and provide information helpful in designing therapeutic strategies for Alzheimer disease.
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PMID:Discovery of a brain promoter from the human transferrin gene and its utilization for development of transgenic mice that express human apolipoprotein E alleles. 861 55

A significant fraction of the beta-amyloid precursor protein is proteolytically processed to yield large secreted forms (sAPP). These proteins have pleiotropic effects which potentially involve control of gene expression. We have investigated the influence of sAPP on the class of transcription factors which bind kappa B enhancer sequences. Transcription dependent on a kappa B element was enhanced by sAPP in several cell lines, as measured by expression of a transfected chloramphenicol acetyltransferase reporter gene. Secreted APP also induced an increase in kappa B DNA-binding activity in hippocampal neurons treated with sAPP. Both effects were mimicked by an analog of cyclic GMP and inhibited by an antagonist of cyclic GMP-dependent protein kinase. Such activation of kappa B-dependent transcription was correlated in two ways with the ability of sAPP to protect neuronal cells against calcium-mediated damage: (1) tumor necrosis factor beta also protected against calcium-mediated insults and induced kappa B-dependent transcription; (2) antisense oligonucleotide-mediated reduction of an endogenous inhibitor of NF-kappa B activated kappa B-binding activity and attenuated calcium-mediated toxicity in both a neuronal cell line and in primary neurons. These findings suggest that a kappa B-binding transcription factor can act as a coordinator of neuroprotective gene expression in response to cytokines.
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PMID:Induction of neuroprotective kappa B-dependent transcription by secreted forms of the Alzheimer's beta-amyloid precursor. 884 19

A change in the regulation of the beta-amyloid precursor protein (beta APP) gene may be a factor for Alzheimer's disease (AD). We have screened a rhesus monkey genomic library and pulled out a approximately 17 kb genomic DNA region which contains the 5'-flanking region (promoter), first exon and intron of the beta APP gene of the Rhesus monkey (rh beta APP). We have partially characterized the genomic clone by selective restriction enzyme digestion followed by Southern blotting against the beta APP gene-specific DNA probes. The identity and authenticity of the different bands of the clone were also confirmed by Southern blot analysis of the rhesus monkey genomic DNA. Functional identification of the genomic fragment was determined by a promoter assay using the chloramphenicol acetyltransferase (CAT) enzyme as a reported gene. Our results showed that a 1.2 kb fragment from the 5'-flanking region of the rh beta APP gene possessed strong promoter activity. The isolation of this genomic clone will enable characterization of the structure and function of the promoter of the rh beta APP gene. Our initial results indicate a high degree of homology between the structure of the rhesus monkey and human beta APP promoter.
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PMID:Isolation of the genomic clone of the rhesus monkey beta-amyloid precursor protein. 984 37

The amyloid precursor protein (APP) has been associated with Alzheimer's disease (AD) because APP is processed into the beta-peptide that accumulates in amyloid plaques, and APP gene mutations can cause early onset AD. Inflammation is also associated with AD as exemplified by increased expression of interleukin-1 (IL-1) in microglia in affected areas of the AD brain. Here we demonstrate that IL-1alpha and IL-1beta increase APP synthesis by up to 6-fold in primary human astrocytes and by 15-fold in human astrocytoma cells without changing the steady-state levels of APP mRNA. A 90-nucleotide sequence in the APP gene 5'-untranslated region (5'-UTR) conferred translational regulation by IL-1alpha and IL-1beta to a chloramphenicol acetyltransferase (CAT) reporter gene. Steady-state levels of transfected APP(5'-UTR)/CAT mRNAs were unchanged, whereas both base-line and IL-1-dependent CAT protein synthesis were increased. This APP mRNA translational enhancer maps from +55 to +144 nucleotides from the 5'-cap site and is homologous to related translational control elements in the 5'-UTR of the light and and heavy ferritin genes. Enhanced translation of APP mRNA provides a mechanism by which IL-1 influences the pathogenesis of AD.
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PMID:Translation of the alzheimer amyloid precursor protein mRNA is up-regulated by interleukin-1 through 5'-untranslated region sequences. 1003 34

The overexpression of the Alzheimer amyloid precursor protein (APP) and its subsequent proteolytic processing may be one of several factors contributing to amyloid beta-peptide (Abeta) deposition in plaques and microvasculature in Alzheimer's disease (AD) brain. Cytokines and growth factors can influence the expression of APP in response to brain injury, but the underlying mechanisms are largely unknown. We examined the mechanisms by which transforming growth factor-beta (TGF-beta) affects the expression of APP in normal human astrocytes. We report that, TGF-beta up-regulated the expression of APP at the transcription level as determined by nuclear run-on experiments. Transient transfection of astrocytes with APP gene promoter (-2832 bp) chloramphenicol acetyltransferase (CAT) reporter constructs led to increased reporter activity upon TGF-beta stimulation. This reporter activity was mainly attributed to the APP proximal domain (-488 bp). The increase in APP gene transcription was associated with significant accumulation of intracellular APP, APP carboxyl terminal derived fragments, and total secreted Abeta. In addition, we observed a significant increase in levels of TGF-beta in Abeta plaques and its immediate vicinity in AD-affected brain relative to controls. These results indicate that high levels of TGF-beta in the cortex, may serve to up-regulate APP synthesis in reactive astrocytes and indirectly contributes to Abeta deposition. Closely related processes may induce cerebrovascular pathology in AD brain.
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PMID:Transcriptional activation and increase in expression of Alzheimer's beta-amyloid precursor protein gene is mediated by TGF-beta in normal human astrocytes. 1209 97

Transforming growth factor-beta-1 (TGF-beta), a key regulator of the brain responses to injury and inflammation, has been implicated in upregulating the expression of the Alzheimer amyloid precursor protein (APP) and Alzheimer's disease (AD) pathogenesis. However, little is known about the mechanisms underlying the effects of TGF-beta on APP expression. Analysis of APP promoter activity upstream of the chloramphenicol acetyltransferase reporter gene in normal human astrocytes (NHAs), revealed that the APP promoter binding beta (APBbeta) site (-93/-82) is responsive to TGF-beta. This site interacts with the zinc finger nuclear factor CTCF, involved in APP transcriptional activity. As determined by gel shift assay, there was no significant difference in the CTCF-APBbeta complex binding activity in the presence or absence of TGF-beta treatment of NHAs. To further investigate the contributions of the CTCF-complex and Smad proteins to the TGF-beta induced APP promoter activity, we examined the distribution of these factors and their DNA binding activity. Interestingly, upon TGF-beta treatment both Smads 3 and 4 were translocated to the nuclei in contrast to Smad 2, which was cytoplasmic. However, CTCF was predominantly localized in the nuclei irrespective of TGF-beta treatment. Gel super shift assay coupled with Western blot analysis showed that Smads 3 and 4 specifically associated with the CTCF-APBbeta complex. In addition, AD brain sections showed increased expression and nuclear localization of Smad 4, which correlated with higher levels of APP and TGF-beta. However, over expression of Smad 4 on its own was not sufficient to affect APP expression. These results demonstrate that TGF-beta activation of Smad protein complexes promotes transcription of the APP gene. Increased synthesis of APP may in part determine Abeta production and deposition in affected AD brain.
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PMID:Transforming growth factor-beta-induced transcription of the Alzheimer beta-amyloid precursor protein gene involves interaction between the CTCF-complex and Smads. 1209 98

Iron-responsive elements (IREs) are the RNA stem loops that control cellular iron homeostasis by regulating ferritin translation and transferrin receptor mRNA stability. We mapped a novel iron-responsive element (IRE-Type II) within the 5'-untranslated region (5'-UTR) of the Alzheimer's amyloid precursor protein (APP) transcript (+51 to +94 from the 5'-cap site). The APP mRNA IRE is located immediately upstream of an interleukin-1 responsive acute box domain (+101 to +146). APP 5'-UTR conferred translation was selectively down-regulated in response to intracellular iron chelation using three separate reporter assays (chloramphenicol acetyltransferase, luciferase, and red fluorescent protein reflecting an inhibition of APP holoprotein translation in response to iron chelation. Iron influx reversed this inhibition. As an internal control to ensure specificity, a viral internal ribosome entry sequence was unresponsive to intracellular iron chelation with desferrioxamine. Using RNA mobility shift assays, the APP 5'-UTRs, encompassing the IRE, bind specifically to recombinant iron-regulatory proteins (IRP) and to IRP from neuroblastoma cell lysates. IRP binding to the APP 5'-UTR is reduced after treatment of cells with desferrioxamine and increased after interleukin-1 stimulation. IRP binding is abrogated when APP cRNA probe is mutated in the core IRE domain (Delta4 bases:Delta83AGAG86). Iron regulation of APP mRNA through the APP 5'-UTR points to a role for iron in the metabolism of APP and confirms that this RNA structure can be a target for the selection of small molecule drugs, such as desferrioxamine (Fe chelator) and clioquinol (Fe, Cu, and Zn chelator), which reduce Abeta peptide burden during Alzheimer's disease.
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PMID:An iron-responsive element type II in the 5'-untranslated region of the Alzheimer's amyloid precursor protein transcript. 1219 35