title: Production of human antibodies by in vitro immunization using a fusion protein containing the transcriptional transactivator of HIV-1.
abstract: Antigen-specific activation of human B cells represents a key step for the production of monoclonal antibodies. Several approaches have been developed over the last thirty years in order to improve the process of lymphocyte activation in vitro. In the present study, we investigated whether the transcriptional transactivator (Tat) of human immunodeficiency virus, which possesses numerous biological activities, is able to trigger antibody secretion when incubated with human peripheral blood mononuclear cells. No such effect was observed when using Tat as a free protein. However, we found a significant IgM antibody production when Tat was previously fused to a double domain, called ZZ, derived from protein A of Staphylococcus aureus. The effect was also observed when the fusion protein, called ZZTat101, was incubated with purified B cells, indicating that the phenomenon does not require T-cell help. Antibody secretion was observed in the absence of cytokines that are usually used during in vitro immunization experiments, indicating that ZZTat101 provides the signals required for the initiation of the immune response. Antibody secretion was observed using a ZZTat mutant, containing only the Tat residues 22 to 57, called ZZTat22-57, indicating that this region is sufficient to initiate the immune response. In contrast, the effect was not found with a ZZTat22-57 mutant devoid of the seven Tat cysteines located between residues 22 and 37, demonstrating that these residues play a crucial role in the phenomenon. Our results pave the way to the development of a new in vitro immunization method based on antigens associated with ZZTat.
authors: CEA, Institut de Biologie et de Technologie de Saclay (iBiTec-S), Service de Pharmacologie et d'immunoanalyse (SPI), 91191 Gif sur Yvette, France.
authors: M
authors: M
authors: Ait Mebarek
authors: A
authors: A
authors: Wijkhuisen
authors: K
authors: K
authors: Adel-Patient
authors: P
authors: P
authors: Lamourette
authors: M
authors: M
authors: Léonetti
authors: H
authors: H
authors: Volland
mesh_terms: D016915:AIDS Vaccines / Q000276:immunology; D000917:Antibody Formation; D001402:B-Lymphocytes / Q000276:immunology; D002478:Cells, Cultured; D015483:HIV Antibodies / Q000276:immunology*; D015658:HIV Infections / Q000276:immunology; D015497:HIV-1 / Q000276:immunology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology; D008213:Lymphocyte Activation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000276:immunology*; D013205:Staphylococcal Protein A / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D013211:Staphylococcus aureus / Q000276:immunology; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D016915:AIDS Vaccines; D015483:HIV Antibodies; D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A; D054322:tat Gene Products, Human Immunodeficiency Virus
keywords: HIV-1 transcriptional transactivator; Human antibodies; In vitro immunization; ZZ domain