Targeting HIV proteins to the major histocompatibility complex class I processing pathway with a novel gp120-anthrax toxin fusion protein
Targeting HIV proteins to the major histocompatibility complex class I processing pathway with a novel gp120-anthrax toxin fusion protein
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DOI：
10.1073/pnas.94.22.12059
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发表时间：
1997-10-28
影响因子：
11.1
通讯作者：
Berzofsky, JA
中科院分区：
文献类型：
--
作者：
Goletz, TJ;Klimpel, KR;Berzofsky, JA
亚单位疫苗的目标是引发CD 8（+）细胞毒性T淋巴细胞（CTL），其挑战是将抗原递送到活细胞的胞质溶胶，在那里它可以被加工以通过主要组织相容性复合体（MHC）I类分子呈递。几种细菌毒素已经进化为有效地将催化蛋白部分递送至真核细胞的胞质溶胶。炭疽致死毒素由两种不同的蛋白质组成，它们联合收割机形成活性毒素，保护性抗原（PA）与细胞结合并有助于将致死因子（LF）递送至细胞胞质溶胶。我们在LF的氨基末端254 aa和HIV-1包膜蛋白的gp 120部分之间构建了遗传融合体，用这种融合蛋白处理的细胞（LF 254-gp 120）在PA存在下有效加工gp 120并呈递HIV-1gp 120 V3特异性CTL识别的表位。当用LF 254-gp 120融合蛋白和易位缺陷的突变PA蛋白处理细胞时，细胞不能呈递表位并且不被特异性CTL裂解，通过显示PA + LF 254 - 1的抗原呈递，证实了进入胞质溶胶和对经典胞质溶胶MHC I类途径的依赖性。用蛋白酶体抑制剂lactacystin阻断gp 120。这些数据证明了LF氨基末端片段将抗原递送至MHC I类途径的能力，并为开发新型T细胞疫苗提供了基础。
A challenge for subunit vaccines whose goal is to elicit CD8(+) cytotoxic T lymphocytes (CTLs) is to deliver the antigen to the cytosol of the living cell, where it can be processed for presentation by major histocompatibility complex (MHC) class I molecules. Several bacterial toxins have evolved to efficiently deliver catalytic protein moieties to the cytosol of eukaryotic cells. Anthrax lethal toxin consists of two distinct proteins that combine to form the active toxin, Protective antigen (PA) binds to cells and is instrumental in delivering lethal factor (LF) to the cell cytosol, To test whether the lethal factor protein could be exploited for delivery of exogenous proteins to the MHC class I processing pathway, we constructed a genetic fusion between the amino-terminal 254 aa of LF and the gp120 portion of the HIV-1 envelope protein, Cells treated with this fusion protein (LF254-gp120) in the presence of PA effectively processed gp120 and presented an epitope recognized by HIV-1 gp120 V3-specific CTL, In contrast, when cells were treated with the LF254-gp120 fusion protein and a mutant PA protein defective for translocation, the cells were not able to present the epitope and were not lysed by the specific CTL, The entry into the cytosol and dependence on the classical cytosolic MHC class I pathway were confirmed by showing that antigen presentation by PA + LF254-gp120 was blocked by the proteasome inhibitor lactacystin. These data demonstrate the ability of the LF amino-terminal fragment to deliver antigens to the MHC class I pathway and provide the basis for the development of novel T cell vaccines.