Wills M R, Carmichael A J, Mynard K, Jin X, Weekes M P, Plachter B, Sissons J G
Department of Medicine, University of Cambridge Clinical School, United Kingdom.
J Virol. 1996 Nov;70(11):7569-79. doi: 10.1128/JVI.70.11.7569-7579.1996.
Cytotoxic T lymphocytes (CTL) appear to play an important role in the control of human cytomegalovirus (HCMV) in the normal virus carrier: previous studies have identified peripheral blood CD8+ CTL specific for the HCMV major immediate-early gene product (IE1) and more recently, by bulk culture and cloning techniques, have identified CTL specific for a structural gene product, the lower matrix protein pp65. In order to determine the relative contributions of CTL which recognize the HCMV proteins IE1, pp65, and glycoprotein B (gB) to the total HCMV-specific CTL response, we have used a limiting-dilution analysis system to quantify HCMV-specific CTL precursors with different specificities, allowing the antigenic specificity of multiple short-term CTL clones to be assessed, in a group of six healthy seropositive donors. All donors showed high frequencies of HCMV-specific major histocompatibility complex-restricted CTL precursors. There was a very high frequency of CTL specific for pp65 (lower matrix protein); IE1-specific CTL were also detectable at lower frequencies in three of five donors, while CTL directed to gB were undetectable. A pp65 gene deletion mutant of HCMV was then used to estimate the contribution of pp65-specific CTL to the total HCMV-specific CTL response; this showed that between 70 and 90% of all CTL recognizing HCMV-infected cells were pp65 specific. Analysis of the peptide specificity of pp65-specific CTL showed that some donors have a highly focused response recognizing a single peptide; the T-cell receptor Vbeta gene usage in these two donors was shown to be remarkably restricted, with over half of the responding CD8+ T cells utilizing a single Vbeta gene rearrangement. Other subjects recognized multiple pp65 peptides: nine new pp65 CTL peptide epitopes were defined, and for five of these the HLA-presenting allele has been identified. All four of the HLA A2 donors tested in this study recognized the same peptide. This apparent domination of the CTL response to HCMV during persistent infection by a single structural protein, irrespective of major histocompatibility complex haplotype, is not clearly described for other persistent virus infections, and the mechanism requires further investigation.
细胞毒性T淋巴细胞（CTL）似乎在正常病毒携带者的人巨细胞病毒（HCMV）控制中发挥重要作用：先前的研究已鉴定出针对HCMV主要立即早期基因产物（IE1）的外周血CD8 + CTL，最近，通过大量培养和克隆技术，已鉴定出针对结构基因产物即低基质蛋白pp65的CTL。为了确定识别HCMV蛋白IE1、pp65和糖蛋白B（gB）的CTL对总HCMV特异性CTL反应的相对贡献，我们使用了有限稀释分析系统来量化具有不同特异性的HCMV特异性CTL前体，从而能够在一组六名健康血清阳性供体中评估多个短期CTL克隆的抗原特异性。所有供体均显示出高频率的HCMV特异性主要组织相容性复合体限制性CTL前体。针对pp65（低基质蛋白）的CTL频率非常高；在五名供体中的三名中也可检测到较低频率的IE1特异性CTL，而未检测到针对gB的CTL。然后使用HCMV的pp65基因缺失突变体来估计pp65特异性CTL对总HCMV特异性CTL反应的贡献；这表明所有识别HCMV感染细胞的CTL中有70%至90%是pp65特异性的。对pp65特异性CTL的肽特异性分析表明，一些供体具有高度集中的反应，识别单一肽；这两名供体中T细胞受体Vβ基因的使用显示出明显受限，超过一半的反应性CD8 + T细胞利用单一Vβ基因重排。其他受试者识别多种pp65肽：定义了九个新的pp65 CTL肽表位，其中五个的HLA呈递等位基因已被鉴定。本研究中测试的所有四名HLA A2供体都识别相同的肽。在持续感染期间，CTL对HCMV的反应明显由单一结构蛋白主导，而与主要组织相容性复合体单倍型无关，其他持续性病毒感染并未明确描述这种情况，其机制需要进一步研究。
