title: Expression and detection of macrophage-tropic HIV-1 gp120 in the brain using conformation-dependent antibodies.
abstract: HIV-1 envelope proteins gp120 and gp41 are likely to play a role in the pathogenesis of HIV-associated neurocognitive disorders. While detection of gp120 in HIV-infected cell cultures is easy, it has not yet been possible to identify gp120 in human or animal brains in situ. The difficulty in detecting gp120 could be due to low expression levels of the protein, to the shedding of gp120 from infected macrophages/microglia, or to the use of inappropriate gp-specific antibodies. We addressed these questions by analyzing the subcellular localization, oligomeric structure, and shedding behavior of gp120 from a macrophage-tropic, CCR5-dependent primary isolate, BX08, expressed by a Semliki Forest virus replicon (SFVenvBX08) in vitro. We used the same SFV system injected in vivo into the rat brain in an attempt to detect gp120 in situ. Our results show that gp120/41 is expressed as monomers, dimers, and trimers in cell culture. Immunocytochemical analysis revealed that intracytoplasmic gp120 can be recognized by an anti-V3 antibody, whereas gp120 at the plasma membrane is detected exclusively by a conformation-dependent antibody. In the rat brain, the SFV vector allows gene expression in neurons from day 3 to day 9 after injection without any apparent brain damage nor reactive astrogliosis. In SFVenvBX08-infected neurons only conformation-dependent antibodies allowed gp120 labeling. These results suggest that previous difficulties in detecting gp120 in brain tissues may be due to the use of antibodies which were unable to recognize gp120 at the plasma membrane.
authors.affiliation: Unité de Virologie Moléculaire, Institut Pasteur, 25, Rue du Dr. Roux, Paris Cedex 15, 75724, France. altmeyer@pasteur.fr
authors.forename: R
authors.initials: R
authors.lastname: Altmeyer
authors.forename: E
authors.initials: E
authors.lastname: Mordelet
authors.forename: M
authors.initials: M
authors.lastname: Girard
authors.forename: C
authors.initials: C
authors.lastname: Vidal
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D015153:Blotting, Western; D001921:Brain / Q000821:virology*; D002460:Cell Line; D005904:Glial Fibrillary Acidic Protein / Q000276:immunology; D015699:HIV Envelope Protein gp120 / Q000032:analysis* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015700:HIV Envelope Protein gp41 / Q000032:analysis / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015497:HIV-1 / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D008264:Macrophages / Q000821:virology*; D051379:Mice; D009474:Neurons / Q000276:immunology / Q000821:virology; D011487:Protein Conformation; D051381:Rats; D012093:Replicon; D012672:Semliki forest virus / Q000235:genetics
chemical_list: D000911:Antibodies, Monoclonal; D005904:Glial Fibrillary Acidic Protein; D015699:HIV Envelope Protein gp120; D015700:HIV Envelope Protein gp41