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)

Human endothelial leukocyte-adhesion molecule 1 (ELAM-1), a cell-surface glycoprotein expressed solely on cytokine-activated endothelial cells, mediates the adhesion of blood neutrophils, memory T-cells and some monocytes. ELAM-1, also known as E-selectin or leukocyte endothelial-cell-adhesion molecule 2, is a member of the lectin/epidermal-growth-factor/complement-regulatory-protein-like cell-adhesion molecule family, which includes structurally related molecules referred to as selectins. They are all involved in cell/cell adhesion, playing roles in leukocyte trafficking which are currently only partially defined. We report here the isolation and characterization of the murine equivalent of human ELAM-1. Murine ELAM-1 is encoded by a single-copy gene, spanning about 13 kb, which is structurally organized into 14 exons and 13 introns; very similar to that of its human counterpart. The exon/intron architecture exactly parallels the domain structure of the encoded protein. A murine ELAM-1-specific cDNA was cloned from heart tissue of an interleukin-1-(IL-1)-treated mouse. Its nucleotide sequence shows an overall similarity of 70% to human ELAM-1 cDNA. Transiently expressed in Cos cells, the encoded protein promotes the adhesion between recombinant cells and both human polymorphic nuclear cells, as well as HL60 cells expressing S-Lewis-x sugar moiety. Northern blot studies revealed by far the highest expression of the murine ELAM-1 gene in heart tissue and only low expression in lung tissue of IL-1-treated mice. Within the promoter, most of the recently identified regulatory elements are conserved. An exception is the nuclear factor (NF) kappa B box sequence, which, in the murine ELAM-1 promoter, does not correspond to the consensus NF kappa B sequence (Lenardo and Baltimore, 1989). Band-shift analyses show no binding to NF kappa B-like proteins. However, fusion of the murine ELAM-1 promoter to a chloramphenicol acetyltransferase reporter confers cytokine-inducible transcription, although at a lower level, when compared to the human ELAM-1 promoter. Our results demonstrate the existence of a murine homologue of the human gene and demonstrate for adhesion functional equivalence between the homologous proteins from the two species. In addition, we provide the first evidence of the utility of the murine model in addressing biological questions about the role which ELAM-1 plays in inflammation.
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PMID:Murine endothelial leukocyte-adhesion molecule 1 is a close structural and functional homologue of the human protein. 137 14

The human 4F2 cell surface antigen is a 120-kilodalton (kDa) disulfide-linked heterodimer which is composed of an 80- to 90-kDa glycosylated heavy chain (4F2HC) and a 35- to 40-kDa nonglycosylated light chain (4F2LC). 4F2 belongs to a family of inducible cell surface molecules which are involved in T-lymphocyte activation and growth. To better understand the molecular mechanism(s) that controls 4F2HC gene expression in both resting and activated T cells, a 4F2HC human genomic clone was isolated and structurally characterized. The 4F2HC gene spans 8 kilobases of chromosome 11 and is composed of nine exons. The 5' upstream region of the gene displays several properties which are characteristic of housekeeping genes. It is G+C rich and hypomethylated in peripheral blood lymphocyte DNA and contains multiple binding sites for the Sp1 transcription factor while lacking TATA or CCAAT sequences. This region of the gene also displays sequence homologies with several other inducible T-cell genes, including the interleukin-2, interleukin-2 receptor alpha chain, dihydrofolate reductase, thymidine kinase, and transferrin receptor genes. A 255-base-pair fragment of the 4F2HC gene which contains 154 base pairs of the 5' flanking sequence was able to efficiently promote expression of the bacterial chloramphenicol acetyltransferase gene in human Jurkat T cells, indicating that it contains promoter or enhancer (or both) sequences. Analyses of chromatin structure in resting and lectin-activated T cells revealed the presence of stable DNase I-hypersensitive sites within both the 5' flanking and intron 1 regions of the 4F2HC gene. Although the 4F2HC gene displayed many of the structural features characteristic of a constitutively expressed gene, lectin-mediated activation of resting peripheral blood T lymphocytes resulted in a dramatic increase in steady-state levels of 4F2HC mRNA.
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PMID:Isolation and structural characterization of the human 4F2 heavy-chain gene, an inducible gene involved in T-lymphocyte activation. 326 70

Colony-stimulating factors (CSFs) are glycoproteins that stimulate the growth of hematopoietic progenitors and enhance the functional activity of mature effector cells. Human granulocyte/macrophage colony-stimulating factor (GM-CSF) is a 22-kDa glycoprotein that stimulates the growth of myeloid and erythroid progenitors in vitro and increases the responsiveness of neutrophils, monocytes, and eosinophils to physiologic stimuli. Elucidation of the cell and tissue sources of CSFs, as well as study of their regulation of expression, is required to understand their role in physiologic and pathophysiologic states. An extensive survey of normal and neoplastic human tissues did not reveal constitutive production of detectable levels of GM-CSF mRNA in any of the 64 samples studied. Antigen- or lectin-activated T lymphocytes have been shown to produce GM-CSF; therefore, to elucidate the genetic sequences required, we constructed recombinant plasmids containing 5' flanking DNA of the GM-CSF gene linked to the marker chloramphenicol acetyltransferase gene. The recombinant constructs were transfected into a human T-cell leukemia virus type I (HTLV)-infected T-lymphoblast cell line that can be stimulated to produce high levels of GM-CSF. We show here that the 5' flanking sequences of the GM-CSF gene can direct increased expression of the chloramphenicol acetyltransferase gene in activated T-lymphoblast cells.
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PMID:Regulation of expression of human granulocyte/macrophage colony-stimulating factor. 349 Jun 69

To provide tools for functional molecular genetics of the protozoan parasite Entamoeba histolytica, we investigated the use of the prokaryotic neomycin phosphotransferase (NEO) gene as a selectable marker for the transfection of the parasite. An Escherichia coli-derived plasmid vector was constructed (pA5'A3'NEO) containing the NEO coding region flanked by untranslated 5' and 3' sequences of an Ent. histolytica actin gene. Preceding experiments had revealed that amoebae are highly sensitive to the neomycin analogue G418 and do not survive in the presence of as little as 2 micrograms/ml. Transfection of circular pA5'A3'NEO via electroporation resulted in Ent. histolytica trophozoites resistant to G418 up to 100 micrograms/ml. DNA and RNA analyses of resistant cells indicated that (i) the transfected DNA was not integrated into the amoeba genome but was segregated episomally, (ii) in the amoebae, the plasmid replicated autonomously, (iii) the copy number of the plasmid and the expression of NEO-specific RNA were proportional to the amount of G418 used for selection, and (iv) under continuous selection, the plasmid was propagated over an observation period of 6 months. Moreover, the plasmid could be recloned into E. coli and was found to be unrearranged. To investigate the use of pA5'A3'NEO to coexpress other genes in Ent. histolytica, a second marker, the prokaryotic chloramphenicol acetyltransferase (CAT) gene under control of an Ent. histolytica lectin gene promoter was introduced into the plasmid. Transfection of the amoebae with this construct also conferred G418 resistance and, in addition, allowed continuous expression of CAT activity in quantities corresponding to the amount of G418 used for selection. When selection was discontinued, transfected plasmids were lost as indicated by an exponential decline of CAT activity in trophozoite extracts.
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PMID:Transfection and continuous expression of heterologous genes in the protozoan parasite Entamoeba histolytica. 756 55

The promoter region driving the gene for the 170-kDa heavy subunit of the Entamoeba histolytica galactose-inhibitable lectin was analysed by transient transfection using the chloramphenicol acetyltransferase gene as reporter. S1 mapping confirmed our previous notion that the promoter is located within a 1.35-kb intergenic sequence preceding the structural lectin gene. Transcripts derived from the chloramphenicol acetyltransferase gene of transfected trophozoites were found to be polyadenylated and the transcriptional start mapped to a position similar to that of the wild-type lectin gene. By deletion analysis the entire promoter was restricted to a fragment covering about 550 bp upstream from the start of transcription. On the other hand, residual promoter activity required a sequence of about 140 bp only, encompassing a newly identified CCAAT-box like element around position -100, as well as the amebic specific TATA-box. This 140-bp fragment as well as a stretch of 15 bp, which is located some 100 nt further upstream, were found to be conserved within the 5' noncoding region of a second E. histolytica lectin gene. Point-mutation analyses indicated that the 15-bp fragment, the likely CCAAT-box, as well as the TATA-box are required for full promoter activity.
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PMID:Analysis of the 170-kDa lectin gene promoter of Entamoeba histolytica. 853 79

Galectin-1 (gal-1), a galactoside-binding lectin, is found in many vertebrate tissues and its expression is regulated during development. We had found that gal-1 expression is increased in F9 murine embryonal carcinoma cells concurrently with induction of differentiation by all-trans retinoic acid (RA). In contrast, gal-1 expression was constitutively high in murine myoblastic C2C12 cells. Therefore, we used these two cell types as models to begin to understand the mechanisms underlying constitutive and RA-induced gal-1 expression. We transfected transiently into F9 cells a series of reporter constructs containing different deletions of the 5' upstream region of the gal-1 gene promoter placed upstream of the chloramphenicol acetyltransferase reporter cDNA and evaluated the activation of transcription by RA treatment. The results indicate that the induction of gal-1 by RA is regulated at least partially at the level of transcription. A strong RA responsiveness region was found within the sequence from -1578 to -1448 upstream of the transcription start site (+1). In contrast, the high constitutive gal-1 expression in C2C12 cells appeared to be mediated by a sequence within the promoter region from -62 to +1, which contains an Sp1 consensus sequence. A gel electrophoretic mobility shift assay indicated that the transcription factor SP1 bound to the gal-1 Sp1 site and mutagenesis of this Sp1 site abolished both the binding of nuclear proteins to the mutated Sp1 site and the high constitutive expression of the gal-1 gene. The results demonstrate that gal-1 expression is cell type-specific and suggest that different factors regulate constitutive and RA-induced gal-1 expression.
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PMID:Differential regulation of constitutive and retinoic acid-induced galectin-1 gene transcription in murine embryonal carcinoma and myoblastic cells. 1076 May 65

We have followed Sp1 expression in primary human T lymphocytes induced, via CD2 plus CD28 costimulation, to sustained proliferation and subsequent return to quiescence. Binding of Sp1 to wheat germ agglutinin lectin was not modified following activation, indicating that the overall glycosylation of the protein was unchanged. Sp1 underwent, instead, a major dephosphorylation that correlated with cyclin A expression and, thus, with cell cycle progression. A similar change was observed in T cells that re-entered cell cycle following secondary interleukin-2 stimulation, as well as in serum-induced proliferating NIH/3T3 fibroblasts. Phosphatase 2A (PP2A) appears involved because 1) treatment of dividing cells with okadaic acid or cantharidin inhibited Sp1 dephosphorylation and 2) PP2A dephosphorylated Sp1 in vitro and strongly interacted with Sp1 in vivo. Sp1 dephosphorylation is likely to increase its transcriptional activity because PP2A overexpression potentiated Sp1 site-driven chloramphenicol acetyltransferase expression in dividing Kit225 T cells and okadaic acid reversed this effect. This increase might be mediated by a stronger affinity of dephosphorylated Sp1 for DNA, as illustrated by the reduced DNA occupancy by hyperphosphorylated Sp factors from cantharidin- or nocodazole-treated cells. Finally, Sp1 dephosphorylation appears to occur throughout cell cycle except for mitosis, a likely common feature to all cycling cells.
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PMID:Sp1 transcriptional activity is up-regulated by phosphatase 2A in dividing T lymphocytes. 1177 71