title: HIV-1 p55Gag encoded in the lysosome-associated membrane protein-1 as a DNA plasmid vaccine chimera is highly expressed, traffics to the major histocompatibility class II compartment, and elicits enhanced immune responses.
abstract: Several genetic vaccines encoding antigen chimeras containing the lysosome-associated membrane protein (LAMP) translocon, transmembrane, and cytoplasmic domain sequences have elicited strong mouse antigen-specific immune responses. The increased immune response is attributed to trafficking of the antigen chimera to the major histocompatibility class II (MHC II) compartment where LAMP is colocalized with MHC II. In this report, we describe a new form of an HIV-1 p55gag DNA vaccine, with the gag sequence incorporated into the complete LAMP cDNA sequence. Gag encoded with the translocon, transmembrane and cytoplasmic lysosomal membrane targeting sequences of LAMP, without the luminal domain, was poorly expressed, did not traffic to lysosomes or MHC II compartments of transfected cells, and elicited a limited immune response from DNA immunized mice. In contrast, addition of the LAMP luminal domain sequence to the construct resulted in a high level of expression of the LAMP/Gag protein chimera in transfected cells that was further increased by including the inverted terminal repeat sequences of the adeno-associated virus to the plasmid vector. This LAMP/Gag chimera with the complete LAMP protein colocalized with endogenous MHC II of transfected cells and elicited strong cellular and humoral immune responses of immunized mice as compared with the response to DNA-encoding native Gag, with a 10-fold increase in CD4+ responses, a 4- to 5-fold increase in CD8+ T-cell responses, and antibody titers of >100,000. These results reveal novel roles of the LAMP luminal domain as a determinant of Gag protein expression, lysosomal trafficking, and possibly of the immune response to Gag.
authors.affiliation: Department of Pharmacology and Molecular Sciences, The Johns Hopkins School of Medicine, Baltimore, Maryland 21205, USA.
authors.forename: Ernesto T A
authors.initials: ET
authors.lastname: Marques
authors.forename: Priya
authors.initials: P
authors.lastname: Chikhlikar
authors.forename: Luciana Barros
authors.initials: LB
authors.lastname: de Arruda
authors.forename: Ihid C
authors.initials: IC
authors.lastname: Leao
authors.forename: Yang
authors.initials: Y
authors.lastname: Lu
authors.forename: Justin
authors.initials: J
authors.lastname: Wong
authors.forename: Juei-Suei
authors.initials: JS
authors.lastname: Chen
authors.forename: Barry
authors.initials: B
authors.lastname: Byrne
authors.forename: J Thomas
authors.initials: JT
authors.lastname: August
mesh_terms: D016475:3T3 Cells; D016915:AIDS Vaccines / Q000276:immunology*; D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D019556:COS Cells; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics / Q000276:immunology*; D004306:Dose-Response Relationship, Immunologic; D058851:GPI-Linked Proteins; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology*; D000949:Histocompatibility Antigens Class II / Q000502:physiology*; D006801:Humans; D007114:Immunization; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D010957:Plasmids; D011498:Protein Precursors / Q000235:genetics / Q000276:immunology*; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000276:immunology*; D019444:Vaccines, DNA / Q000276:immunology*
chemical_list: D016915:AIDS Vaccines; D015816:Cell Adhesion Molecules, Neuronal; D058851:GPI-Linked Proteins; D015683:Gene Products, gag; D000949:Histocompatibility Antigens Class II; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; C095362:limbic system-associated membrane protein; C059767:p55 gag precursor protein, Human immunodeficiency virus 1