Immunological impact of Tetherin retrovirus restriction

描述（由申请人提供）：免疫系统的先天臂可以关键地塑造针对病原体的适应性免疫应答。因此，先天感应机制已被广泛研究，用于合理的佐剂设计。相比之下，对于能够直接抑制病毒复制的“限制因子”基因的免疫学影响知之甚少。Tetherin是被大流行性HIV-1的Vpu抵消以促进病毒体释放的蛋白质。在不存在Vpu的情况下，Tetherin将病毒体“拴系”在细胞表面上。随后，这些拴系的病毒体通过拴系蛋白介导的内吞作用被带回感染的细胞中。Tetherin的免疫生物学仍然未知，体外研究引起了关于它是否作为逆转录病毒限制因子的争议。因此，分离基因进行对照研究的体内研究可能有助于提高对Tetherin功能的认识并解决差异。此外，研究体内Tetherin的生物学可能会受益于破坏其神秘的内吞功能。最近，我们鉴定了具有导致内吞缺陷的Tetherin单核苷酸多态性（SNP）的近交系小鼠品系（Barrett等人PLOS Pathogens 2012）。在已发表和未发表的遗传回交研究中，我们发现这种Tetherin内吞作用SNP对Friend逆转录病毒（FV）感染的影响取决于主要组织相容性复合体（MHC）基因座。此外，表达Tetherin的MHC II类+抗原呈递细胞（APC）比Tetherin缺陷型APC更大程度地刺激FV特异性CD 4 + T细胞杂交瘤。因此，Tetherin介导的病毒粒子内吞作用在APC中可能增加用于MHC II类呈递的病毒肽库，并增强逆转录病毒特异性CD 4 + T细胞应答。我们建议使用两个互补的系统来测试这一假设。在具体目标1中，我们将评估表达野生型、敲除型和内吞缺陷型Tetherin的APC在FV感染模型中刺激CD 4 + T细胞的能力。由于Tetherin也可能影响对MHC I类的交叉呈递，我们还将研究CD 8 + T细胞应答。在Specific Aim 2中，我们将测试通过破坏性Vpu突变实现Tetherin活性是否会使HIV-1感染的靶细胞更容易受到体外HIV-1特异性CD 4 + T细胞克隆的细胞溶解和抑制作用的影响。来自拟议的体内免疫学研究和HIV-1体外研究的一致结果可能为利用Tetherin生物学用于逆转录病毒免疫治疗和T细胞疫苗提供见解。

DESCRIPTION (provided by applicant): The innate arm of the immune system can critically shape the adaptive immune response against pathogens. Thus, innate sensing mechanisms have been studied extensively for rational adjuvant design. In contrast, much less is known about the immunological impact of 'restriction factor' genes that could directly inhibit virus replication. Tetherin is the protein counteracted by Vpu of pandemic HIV-1 to promote virion release. In the absence of Vpu, Tetherin 'tethers' virions on the cell surface. Subsequently, these tethered virions are brought back into the infected cell through Tetherin-mediated endocytosis. The immunobiology of Tetherin remains unknown, and in vitro studies raised controversies on whether it acts as a retrovirus restriction factor. Thus, in vivo studies that isolate the gene for controlled study may help advance insights and resolve discrepancies on Tetherin function. In addition, investigating the biology of Tetherin in vivo may benefit from disrupting its mysterious endocytic function. Recently, we identified an inbred mouse strain that harbored a Tetherin single nucleotide polymorphism (SNP) that resulted in an endocytosis defect (Barrett et al. PLOS Pathogens 2012). In published and unpublished genetic backcrossing studies, we discovered that the impact of this Tetherin endocytosis SNP on Friend retrovirus (FV) infection is dependent on the Major Histocompatibility Complex (MHC) locus. Moreover, MHC Class II+ antigen-presenting cells (APCs) expressing Tetherin stimulated an FV- specific CD4+ T cell hybridoma to a greater extent than Tetherin-deficient APCs. Thus, Tetherin-mediated virion endocytosis in APCs may increase the viral peptide pool for MHC Class II presentation and augment the retrovirus-specific CD4+ T cell response. We propose to test this hypothesis using two complementary systems. In Specific Aim 1, we will evaluate the ability of APCs expressing wild-type, knock- out and endocytosis-defective Tetherin to stimulate CD4+ T cells in the FV infection model. Since Tetherin may also impact cross-presentation to MHC Class I, we will also investigate the CD8+ T cell response. In Specific Aim 2, we will test if enabling Tetherin activity through a disruptive Vpu mutation would render the HIV-1 infected target cells more susceptible to the cytolytic and suppressive effects of HIV-1 specific CD4+ T cell clones in vitro. Concordant results from the proposed in vivo immunologic studies and HIV-1 in vitro studies may provide insights in harnessing Tetherin biology for retroviral immunotherapeutics and T-cell vaccines.