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
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
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DOI：
10.1074/jbc.m303336200
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发表时间：
2003-09-26
影响因子：
4.8
通讯作者：
August, JT
中科院分区：
文献类型：
--
作者：
Marques, ETA;Chikhlikar, P;August, JT
编码含有溶酶体相关膜蛋白（LAMP）易位子、跨膜和胞质结构域序列的抗原嵌合体的几种基因疫苗已引起强烈的小鼠抗原特异性免疫应答。增加的免疫应答归因于抗原嵌合体向主要组织相容性II类（MHC II）区室的运输，其中LAMP与MHC II共定位。在这份报告中，我们描述了一种新形式的HIV-1 p55 gag DNA疫苗，与gag序列纳入完整的LAMP cDNA序列。用LAMP的转位子、跨膜和细胞质溶酶体膜靶向序列编码的Gag，没有管腔结构域，表达差，不运输到转染细胞的溶酶体或MHC II区室，并且从DNA免疫的小鼠中引起有限的免疫应答。相反，向构建体中添加LAMP管腔结构域序列导致转染细胞中LAMP/ Gag蛋白嵌合体的高水平表达，其通过向质粒载体中包含腺相关病毒的反向末端重复序列而进一步增加。这种具有完整LAMP蛋白的LAMP/ Gag嵌合体与转染细胞的内源性MHC II共定位，并且与对编码DNA的天然Gag的应答相比，引起免疫小鼠的强细胞和体液免疫应答，其中CD 4（+）应答增加10倍，CD 8（+）T细胞应答增加4至5倍，并且抗体滴度> 100，000。这些结果揭示了LAMP管腔结构域作为Gag蛋白表达、溶酶体运输和可能的对Gag的免疫应答的决定因素的新作用。
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.