Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We demonstrated different transduction efficiency in several major organs of the immature (newborn) versus mature (adult) mice using adenoviral recombinants containing expression cassettes for either firefly luciferase or bacterial beta-galactosidase reporter genes. The studied tissues included skeletal muscle, heart, brain, lung, kidney, and liver. The transduction efficiency in all tissues, especially skeletal muscle, was significantly less in adults than in newborns, with two exceptions. In the heart, transduction efficiency was the same in newborns and adults, while in brain, it was greater in the adult than in the newborn. The cited differences in transduction efficiencies between newborn and adult tissues applied approximately equally to both reporter genes. The alpha v integrin level showed the same trend as the transduction efficiency in all tissues, except the heart. Polymerase chain reaction showed a specific adenoviral product in proportion to the reporter gene expression in muscle, heart, and brain. The results of this study should be considered in designing gene therapy strategies in genetic diseases.
Exp Mol Pathol 1995 Apr
PMID:Differential short-term transduction efficiency of adult versus newborn mouse tissues by adenoviral recombinants. 854 97

The insulin-like growth factor-II/cation-independent mannose 6-phosphate receptor (IGF-II/MPR) is a multifunctional protein that binds IGF-II and ligands containing a mannose 6-phosphate recognition marker. Recent studies have shown that this receptor plays a critical role in mammalian development, and that its expression is controlled by both epigenetic and tissue-specific factors. Our laboratory has cloned the 93-kilobase mouse gene and characterized its 48 exons. In this report we describe the structure and function of the IGF-II/MPR gene promoter. To study promoter function, a series of chimeric plasmids linking different segments of IGF-II/MPR 5' flanking DNA to the reporter gene, firefly luciferase, were transiently transfected into HepG2 and C3H 10T1/2 cells. Promoter activity was orientation-specific and was maximal (550- to 4250-fold above promoterless control) with a plasmid containing 266 base pairs (bp) of IGF-II/MPR DNA. The fusion gene accurately directed transcription as measured by ribonuclease protection assay using RNA extracted from transfected cells. DNA-protein binding studies by in vitro DNase I footprinting revealed an extended 54-bp footprint within the proximal promoter that contained two E-boxes and potential binding sites for transcription factors Sp1, NGF-IA, and related proteins. Gel mobility shift experiments with double-stranded oligonucleotides containing this region gave rise to several specific DNA-protein complexes, and the addition of specific antibodies indicated that proteins antigenically related to Sp1 and c-Myc were components of one or more of these bands. Deletion of this 54-bp segment led to an 8-fold decline in promoter activity, and its transfer to a heterologous promoter stimulated gene expression by nearly 7-fold. Mutational analyses indicated that each E box contributed to more than half of the enhancer's activity. These results define a strong minimal IGF-II/MPR promoter of no more than 266 bp and identify a 54-bp enhancer within this promoter fragment. Our observations thus represent a first step toward characterizing the developmental, epigenetic, and tissue-specific factors that control IGF-II/MPR gene expression.
Mol Endocrinol 1995 Nov
PMID:Control of insulin-like growth factor-II/mannose 6-phosphate receptor gene transcription by proximal promoter elements. 858 25

Calreticulin is a new human rheumatic disease-associated autoantigen that plays a multifaceted role in cell biology. In earlier studies, this protein was shown to share an intimate relationship with the Ro/SS-A autoantigen complex, although the nature of this association continues to be debated. Since modulation of the Ro/SS-A autoantigen in epidermal keratinocytes has been implicated in the pathogenesis of subacute cutaneous lupus erythematosus and neonatal lupus erythematosus, we have begun to examine the transcriptional regulation of calreticulin. A 504 bp calreticulin promoter fragment was subcloned into a reporter gene plasmid containing firefly luciferase. Calcium ionophore, heat shock, and heavy metals such as zinc and cadmium were consistently found to increase calreticulin transcriptional activities in A431 cells (a human epidermoid squamous carcinoma cell line) under transient transfection conditions. These studies suggest that (a) calreticulin is regulated at the transcriptional level, and (b) calreticulin, like some other LE-related autoantigens, appears to function as a heat shock/stress-response gene.
Mol Immunol
PMID:Calreticulin is transcriptionally upregulated by heat shock, calcium and heavy metals. 867 89

Expression of the firefly luciferase gene under the control of viral or fish promoters was observed in fish tissue after direct DNA injection of plasmid DNA. Plasmid DNA containing the firefly luciferase gene was injected into the skeletal muscle of rainbow trout (Oncorhynchus mykiss), and levels of luciferase activity were found to be dependent on the controlling promoter and the amount of injected DNA. Plasmids using the cytomegalovirus immediate early promoter (CMV-IEP) consistently produced the highest levels of luciferase activity. Maximal activity was observed five to seven days postinjection with 50 micrograms of DNA. This activity persisted in the tissues for as long as 115 days postinjection. When the DNA was examined up to two months postinjection, the predominant form was unreplicated, unintegrated DNA in linear and relaxed circular conformation. Expression of injected DNA was found predominantly within muscle cells along the injection path and in scattered muscle cells anterior to the injection site.
Mol Mar Biol Biotechnol 1996 Jun
PMID:Gene expression in rainbow trout (Oncorhynchus mykiss) following intramuscular injection of DNA. 868 May 23

The effects of 3' untranslated regions (UTR) and intergenic regions (INT), from various Trypanosoma cruzi stage-specific and constitutive genes, on the expression of the reporter firefly luciferase gene (luc), were studied using stable episomal transformation. The 3' UTR influenced luciferase expression by changing the steady-state level and/or the translation efficiency of luc mRNA. Glycoprotein 72 gene (gp72), glycoprotein 85 gene (gp85) or amastin gene (ama) 3' UTR decreased the luc mRNA level 6- to 14-fold, compared to the glyceraldehyde 1-phosphate dehydrogenase gene (gapdh) 3' UTR, in epimastigotes. Luciferase activity decreased in parallel with the luc mRNA level in transformants utilizing the gp85 or ama 3' UTR, whereas luc mRNA containing the gp72 3' UTR showed approximately 5-fold higher translation efficiency than luc mRNA containing a minimal 3' UTR. In amastigotes, the inhibitory effect of the ama 3' UTR observed in other life cycle stages was abolished and luciferase expression was stimulated 16-fold. The overall stage-specific difference mediated by the ama 3' UTR, between epimastigotes and amastigotes, was approximately 100-fold. INT, which was expected to influence polyadenylation efficiency, of gapdh, gp72, or heat shock protein 60 gene inserted after gapdh 3' UTR increased luc mRNA 2- to 8-fold, whereas gp85 INT slightly decreased luc mRNA. By separating effects attributable to the 3' UTR and INT, this study shows the effects of 3' UTR on RNA levels and translational efficiency in T. cruzi.
Mol Biochem Parasitol 1995 Dec
PMID:Effects of 3' untranslated and intergenic regions on gene expression in Trypanosoma cruzi. 872 Jan 75

GroEL and DnaK with their cofactors constitute the major chaperone systems promoting protein folding in the Escherichia coli cytosol. The ability of GroEL to bind and promote folding of a substrate released from DnaK led to the proposal that the DnaK and GroEL systems act successively along a protein folding pathway. Here we have investigated the role of both systems in preventing aggregation and assisting refolding of firefly luciferase denatured by guanidinium chloride and heat. We find that DnaK and GroEL compete with each other for binding to non-native luciferase. Addition of ATP and co-operating proteins results in release of luciferase from either chaperone in a non-native conformation. Only a small fraction of luciferase molecules released from GroEL can reach the native state. Instead, the released luciferase must bind repeatedly to the DnaK system, and only then is it able to fold to the native state. Thus, during a folding reaction, DnaK and GroEL do not obligatorily act in succession by promoting earlier and later protein folding steps, respectively. Rather, the two chaperone systems and perhaps others can form a lateral network of co-operating proteins. This chaperone network is proposed to be of particular importance for the assisted refolding of proteins that are unfolded by stress treatment such as heat shock and whose size is too large to allow folding inside the substrate binding cavity of the GroEL ring underneath GroES.
J Mol Biol 1996 Aug 23
PMID:Substrate shuttling between the DnaK and GroEL systems indicates a chaperone network promoting protein folding. 878 Jul 75

HSP 70.1 is one of the first genes to be expressed in the mouse embryo at the time of zygotic genome activation. We studied the regulation of this gene, using a transgene associating HSP 70.1 promoter and the firefly luciferase reporter gene, which allows the precise quantification of HSP 70.1 level of expression on individual embryos. In the present work, we show first that the level of HSP 70.1 expression at the two-cell stage is significantly higher (around two-fold) in embryos whose maternal cytoplasm is from C3H strain than with BALB/c strain. We verified that this difference is not an artefact of the use of transgenic embryos, of the time of first cleavage, or of in vitro culture. This regulation of HSP 70.1 level of expression is controlled by strain-specific maternal modifiers and is independent of replication, syngamy, and mitosis. Following nuclear transfer, reactivation of HSP 70.1 is also subjected to the same epigenetic influence. Only the strain-of-origin of the recipient cytoplast modulates the level of HSP 70.1 reprogrammation; the origin of donor nucleus is not significant, demonstrating the reversibility of this strain effect. These results point out the importance of the quality of recipient cytoplast in the intensity of gene reprogrammation, which may be of importance for nuclear transfer efficiency.
Mol Reprod Dev 1996 Aug
PMID:Quantitative control of gene expression by nucleocytoplasmic interactions in early mouse embryos: consequence for reprogrammation by nuclear transfer. 884 84

There are two types of transcripts for the human A1, adenosine receptor. They are expressed in a tissue-specific manner in human tissues and contain distinct exons. Previously, it had appeared that the two transcripts may have occurred through alternative splicing. The transcript beta has two upstream AUG codons, which in transiently transfected COS-7 cells leads to a reduced level of receptor expression. When genomic sequence including sequences 5' to transcriptional start site, exon 1A, intron 1A, exon 1B, intron 1B, exon 2, and coding sequence was inserted into an expression vector (pCMV5/huA1), the resulting transcripts had the same overall structure as the transcripts present in human tissues. Primer extension and 5' rapid amplification of cDNA ends of mRNA from transfected cells revealed the transcription start sites for these two transcripts occurred in what previously had been termed introns. These results were confirmed with similar analysis of mRNA derived from human tissues. Two nonconsensus putative TATA boxes (TTAAGA and TTTAAA) are located upstream of the transcription start sites for transcripts alpha and beta. When the TATA boxes and their flanking sequences were fused to a firefly luciferase gene containing promoterless vector, both demonstrated strong promoter activity in Chinese hamster ovary cells. Promoter A directs the synthesis of transcript alpha, and promoter B directs the synthesis of transcript beta. Promoter A contains a series of AGG elements between the putative TATA box and the transcription start, which accounts for a major portion of the promoter activity based on deletion and mutation analysis. In general, promoter A is more active than promoter B in transfected cells. The nonconsensus TATA box in promoter B plays a more important role in promoter activity than the TATA box in promoter A. The human A1 adenosine receptor gene appears to use two separate promoters to direct synthesis of distinct transcripts, which can then regulate the relative abundance of A1 adenosine receptor in tissues. We have redefined the human A1 adenosine receptor gene structure based on these new data.
Mol Pharmacol 1995 Dec
PMID:Separate promoters in the human A1 adenosine receptor gene direct the synthesis of distinct messenger RNAs that regulate receptor abundance. 884 13

The flanking sequences of several genes have been shown to direct a position independent expression of transgenes. Attempts to completely identify the insulating sequences have failed so far. Some of these sequences contain a matrix attached region (MAR) located in the flanking part of the genes. This article will show that the MARs in cultured cells located in the 3' OH region of the human apolipoprotein B100 (Apo B100) and within the SV40 genome were unable to stimulate and insultate transgene expression directed by the promoters from a rabbit whey acidic protein (WAP) gene or from human cytomegalovirus (hCMV) early genes. In transgenic mice, the MAR from the Apo B100 and SV40 genes did not enhance the expression of a transgene containing the rabbit whey acid protein (WAP) promotor, the late gene SV40 intron (VP1 intron), the bovine growth hormone (bGH) cDNA and the SV40 late gene terminator. This construct was even toxic for embryos. Similarly, the specialized chromatin structure (SCS) from the Drosophila 87A7 HSP70 gene reduced chloramphenicol acetyl transferase (CAT) activity when added between a cytomegalovirus (CMV) enhancer and a Herpes simplex thymidine kinase (TK) gene promoter. This inhibitory action was almost complete when a second SCS sequence was added before the CMV enhancer. Sequences from the firefly luciferase and from the human gene cathepsin D cDNA used as control unexpectedly showed a similar inhibitory effect when added to the CMVTKCAT construct instead of SCS. When added before the CMV enhancer and after the transcription terminator in the CMVTKCAT construct, the SCS sequence was unable to insulate the integrated gene as seen by the fact that the level of CAT in cell extracts were by no means correlated with the number of copies in individual clones. From these data, it is concluded that i) a MAR containing the canonical AT rich sequences does not amplify the expression of all gene constructs ii) At rich MAR sequences do not have per se an insulating effect iii) Drosophila SCS from the 87A7 HSP70 gene has no insulating effect in all gene constructs (at least in mammalian cells) iv) and the addition of a DNA fragment between an enhancer and a promoter in a gene construct cannot be used as a reliable test to evaluate its insulating property.
Mol Biol Rep
PMID:The effect of matrix attached regions (MAR) and specialized chromatin structure (SCS) on the expression of gene constructs in cultured cells and in transgenic mice. 885 71

Herpes simplex virus-based amplicon vectors have been used for gene transfer into cultured neurons and the adult CNS. Since constitutive expression of a foreign gene or overexpression of an endogenous gene may have deleterious effects, the ability to control temporal expression would be advantageous. To achieve inducible gene expression, we have incorporated the tetracycline-responsive promoter system into amplicon vectors and showed, both in vitro and in vivo, that expression can be modulated by tetracycline. Using the firefly luciferase as the reporter gene, maximal repression by tetracycline in hippocampal cultures was about 50-fold. Withdrawal of tetracycline derepressed gene expression, reaching maximal levels within 10-12 h. In contrast, addition of tetracycline to cultures without prior tetracycline exposure inhibited gene expression rapidly; luciferase activity was reduced to less than 8% within 24 h. In adult rat hippocampus, vectors expressing luciferase or the Escherichia coli lacZ were repressed by tetracycline 9- and 60-fold, respectively. Maximum gene expression from the vectors occurred 2-3 days post-infection and declined thereafter. Such decline impeded further induction of expression by withdrawing tetracycline. This study demonstrates the feasibility of incorporating a powerful inducible promoter system into HSV vectors. The development of such an inducible viral vector system for gene transfer into the adult CNS might prove to be of experimental and therapeutic value.
Brain Res Mol Brain Res 1996 Sep 05
PMID:Inducible gene expression from defective herpes simplex virus vectors using the tetracycline-responsive promoter system. 888 53


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