RESUMO
To investigate parvoviral interference with human immunodeficiency virus type 1 (HIV-1) in human cells that are normally susceptible to HIV-1 infection, nonstructural (NS) proteins of the parvoviruses H-1 virus and minute virus of mice were studied for their effect on the activity of the HIV-1 promoter in a variety of CD4+ cells. Transient cotransfection assays revealed a reduced HIV-1 promoter activity in the presence of parvoviral NS proteins. Stimulation of the HIV-1 promoter by phorbol 12-myristate 13-acetate (PMA) led to an increase in its sensitivity to NS-induced suppression. The inhibitory effect of NS polypeptides depended, at least in part, on the presence of the NF kappa B motifs of the HIV-1 long terminal repeat, suggesting an interaction of the parvoviral products with PMA-inducible cellular factors binding to these elements of the HIV-1 promoter.
Assuntos
Regulação Viral da Expressão Gênica/efeitos dos fármacos , HIV-1/crescimento & desenvolvimento , Parvovirus/crescimento & desenvolvimento , Interferência Viral/genética , Proteínas não Estruturais Virais/farmacologia , Animais , Células Cultivadas , Genes Reporter , Repetição Terminal Longa de HIV/genética , HIV-1/genética , Humanos , Luciferases/biossíntese , Luciferases/genética , Vírus Miúdo do Camundongo/genética , Vírus Miúdo do Camundongo/crescimento & desenvolvimento , NF-kappa B/metabolismo , Parvovirus/genética , Regiões Promotoras Genéticas/genética , Ratos , Sequências Reguladoras de Ácido Nucleico/genética , Supressão Genética , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
Recent findings have indicated that megakaryocytes may be susceptible to human immunodeficiency virus (HIV) infection, suggesting a potential role for megakaryocytes as viral reservoirs in HIV-infected patients. We report that the megakaryocytic cell line Dami could be productively infected with the HTLV III-B strain of HIV-1, in 26 different experiments (results of 16 experiments are reported); productive infection lasted up to 30 weeks. Despite a lack of detectable surface expression of the CD4 molecule and very low levels of CD4 mRNA, between 40% and 60% of megakaryocytic cells produced viral proteins after contact with HIV-1. Neither cytopathogenic effects nor syncytial formation was observed. Production of high levels of functional viral particles was indicated by analysis of p24 protein levels, reverse transcriptase activity, ultrastructural studies, and the capacity of supernatants from infected Dami cells to infect the Molt-4 T-lymphocytic cell line. HIV-1 RNA and protein levels in infected Dami cells were enhanced by treatment with tumor necrosis factor-alpha (TNF-alpha), and decreased by treatment with interferon-alpha (IFN-alpha) and IFN-gamma. Transient transfection of the megakaryocytic cells with various constructs of the HIV-1 promoter (LTR) linked to the luciferase reporter gene suggested that the effect of TNF-alpha was related, as in monocytic and T-cell lines, to transactivation of the enhancer region of the HIV-1 LTR. These findings indicate that signals provided by the immune system may modulate HIV-1 expression in cells of the megakaryocytic lineage.
Assuntos
Citocinas/farmacologia , HIV-1/fisiologia , Megacariócitos/citologia , Replicação Viral/fisiologia , Linhagem Celular , Proteína do Núcleo p24 do HIV/análise , Proteína do Núcleo p24 do HIV/genética , Repetição Terminal Longa de HIV , HIV-1/efeitos dos fármacos , HIV-1/genética , Humanos , Técnicas Imunoenzimáticas , Cinética , Megacariócitos/microbiologia , Megacariócitos/ultraestrutura , Microscopia Eletrônica , RNA Viral/genética , RNA Viral/metabolismo , Ativação Transcricional/efeitos dos fármacos , Transfecção , Fator de Necrose Tumoral alfa/farmacologia , Replicação Viral/efeitos dos fármacosRESUMO
IgE FcR (FcERII) on human eosinophils was characterized and compared with FcERII present on B cells (CD23). Two mAb, BB10 (anti-eosinophil FcERII) and 135 (anti-CD23), bound to the major component of FcERII at 45,000 to 50,000 Mr, both on purified hypodense eosinophils and on a B cell line (WIL-2WT). The specific ligand, human myeloma IgE, was able to bind to the molecules immunoprecipitated by BB10. A cross-reactivity between BB10 and a mAb anti-Leishmania gp63, which is a "fibronectin (Fn)-like" molecule, containing the L-arginine-L-glycyl-L-aspartyl (RGD) cell attachment domain indicated the presence of such a sequence in the common structure present on eosinophil and B cell FcERII. The synthetic tetrapeptide RGDS as well as its inverted sequence (SDGR) reduced the binding of BB10 and anti-Fn mAb to eosinophils and B cells. Flow microfluorometry analysis revealed a variable binding of BB10 and anti-Fn mAb to eosinophils purified from different patients, results compatible with recent findings on the inducibility of FcERIIb. The significant inhibition of IgE-dependent cytotoxicity against parasite targets by preincubation of eosinophils with BB10, anti-Fn and anti-CD23 mAb, with anti-RGDS polyclonal antibodies or with the SDGR peptide suggested the requirement of this cell adhesion sequence for the function of low affinity FcERII. The presence of such a sequence in the C-terminal domain of B cell FcERII raised the possibility of its role in B cell adhesion or B cell growth.