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)

The binding of human immunodeficiency virus type 1 (HIV-1) gp120env to CD4 is the first event leading to infection and represents an important target for possible therapeutic intervention. To provide a tool for screening and quantitation of the effects of drugs inhibiting the Env-CD4 interaction, we developed a simple, fast and quantitative bioassay measuring the fusion between two cell lines generated by stable transfection: one expressing high levels of HIV-1 proteins but no infectious virus (HL2/3), and the other expressing the CD4 receptor and containing an inducible chloramphenicol acetyltransferase (CAT) gene linked to the HIV-1 long terminal repeat (HLCD4-CAT). Upon cocultivation of HL2/3 and HLCD4-CAT cells, efficient cell fusion is observed within 8 h. The efficiency of fusion can be evaluated visually and quantitated by measuring CAT enzyme. This novel bioassay allows testing for drugs capable of interfering with the CD4-Env interaction. HL2/3 cell line secretes gp120env in the medium and can be used for the production of Env protein.
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PMID:A bioassay for HIV-1 based on Env-CD4 interaction. 207 9

We constructed a human immunodeficiency virus (HIV) trans-activator cDNA (tat) encoding the N-terminal 76 amino acids of the viral trans-activator followed by two additional amino acids (val and pro). This cDNA encoded a functional trans-activator (TAT) as shown by cotransfection into murine cells with a HIV promoter-chloramphenicol acetyltransferase DNA construct. The tat cDNA was cloned into an avian retroviral expression vector, a modified spleen necrosis virus (SNV), and high-titer infectious stocks of recombinant virus (SNV-tat) were recovered from dog cells. Hybridization analyses indicated that SNV-tat was stably propagated in these cells for months. We also prepared recombinant cells that stably carry reporter genes, either a human gene encoding a soluble CD4 receptor (sCD4) or the human preprorenin gene, under the transcriptional control of the HIV promoter. Medium obtained from these cell cultures after infection with control viruses or an SNV carrying an antisense tat contained only low background levels of sCD4 or prorenin (HRN) as determined by specific immunoassays (1-10 ng protein per 10(6) cells per ml medium). In contrast, cells infected with SNV carrying tat in the transcriptional sense orientation secreted 75 +/- 7 ng sCD4 and 73 +/- 4 ng HRN per 10(6) cells per ml medium. Moreover, these proteins were constitutively secreted at these levels during months of subculturing. The data indicate that sCD4 and HRN are secreted from these cells because of a TAT-mediated trans-activation of the HIV reporter gene DNA and/or RNA. This combination of recombinant cells, SNV-tat, and specific immunoassays provide a rapid, quantitative, and safe bioassay to seek inhibitors of TAT.
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PMID:A rapid, quantitative bioassay based on the human immunodeficiency virus trans-activator. 259 May 54

Several laboratories have presented evidence that HIV can productively infect CD4- cell lines. However, this data could be challenged on the basis that the target cells may express low levels of the CD4 receptor. In addition, it could be argued that assays might be detecting residual virus. In the case of cell-mediated infection, it is possible that virus detected in assays could be secreted from HIV-infected donor cells rather than the target CD4- cells. In this report we describe a CD4- epithelial cell line which has been transfected with a plasmid containing the chloramphenicol acetyltransferase (CAT) gene ligated to the HIV LTR. CAT-ELISA and immunocytochemistry indicate that target cells synthesize CAT after exposure to HIV-infected primary activated peripheral blood mononuclear cells (PBMC). Results correlate very well with p24 ELISA assays. Infection of epithelia by primary NSI strains of HIV can be blocked by patient antisera or by certain sulfated polysaccharides. Since the CAT assay is not dependent on virus production, the data reported here confirm that CD4- epithelial cells derived from the human cervix can be productively infected by HIV. The observations also support the theory that sexual transmission of HIV could be initiated by infection of genital tract epithelia. Furthermore, the findings support the suppositions that sexual transmission of HIV could be prevented by antibodies to HIV or alternately by a topical formulation containing certain sulfated polysaccharides.
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PMID:CAT-transfected epithelial cells provide evidence for a CD4 independent pathway of HIV infection. 1021 19