Lewicki H, Tishon A, Borrow P, Evans C F, Gairin J E, Hahn K M, Jewell D A, Wilson I A, Oldstone M B
Department of Neuropharmacology, Scripps Research Institute, La Jolla, California 92037, USA.
Virology. 1995 Jun 20;210(1):29-40. doi: 10.1006/viro.1995.1314.
Cytotoxic T lymphocytes (CTL) play a pivotal role in preventing persistent viral infections and aborting acute infections. H-2Db-restricted CTL optimally recognize a specific peptide of 9 to 11 amino acids (aa) derived from a viral protein and held in place (restricted) by a MHC class I glycoprotein on the surfaces of infected cells. Only three peptide sequences with the appropriate Db motif from lymphocytic choriomeningitis virus Armstrong strain (LCMV) are known to be presented to CTL by H-2Db molecules; they are from the glycoproteins (GP), residues 33-41 KAVYNFATC (GP1) and 276-286 SGVENPGGYCL (GP2), and the nucleoprotein (NP), 396-404 FQPQNGQFI. Incubation of virally infected H-2b cells with CTL clones that recognize only GP1, GP2, or NP leads to the selection of viral variants which upon infecting cells bearing H-2b molecules, escape recognition by CTL of the appropriate specificity. Nucleic acid sequencing showed a single mutation in GP1 (aa 38 F-->L), GP2 (aa 282 G-->D), or NP (aa 403 F-->L) in the variant viruses. When wild-type (wt) LCMV peptides and the three variant peptides (GP1, GP2, NP) were synthesized and subjected to a competitive inhibition binding assay, no differences in binding affinity for H-2Db were found between the wt and variant peptides. Uninfected cells coated with the wt peptide were recognized and lysed by the appropriate CTL clone or by in vivo-primed bulk CTL, but similar targets coated with the GP1, GP2, or NP variant peptides were not. This result, coupled with computer graphic analysis of these variant peptides with the recently solved three-dimensional structure for the Db MHC class I molecule, placed the side chain of the mutated residues on the outer surface of the MHC-peptide complex and accessible to the T cell receptor. Ala substitution at GP residue 38 or 282 or at NP 403 also abrogated CTL recognition and lysis. Inoculation of any one of the mutated viral variants into mice produced an effective CTL response to the other two nonmutated GP or NP peptides, suggesting that production of biologically relevant CTL escape virus variants in vivo requires selection of mutations in more than one and likely all the CTL epitopes, a low probability event.
细胞毒性T淋巴细胞（CTL）在预防持续性病毒感染和终止急性感染中起关键作用。H-2Db限制性CTL能最佳地识别源自病毒蛋白的9至11个氨基酸（aa）的特定肽段，该肽段由感染细胞表面的MHC I类糖蛋白固定在位（受限）。已知淋巴细胞性脉络丛脑膜炎病毒阿姆斯特朗株（LCMV）仅有三个具有合适Db基序的肽序列可由H-2Db分子呈递给CTL；它们分别来自糖蛋白（GP），残基33 - 41 KAVYNFATC（GP1）和276 - 286 SGVENPGGYCL（GP2），以及核蛋白（NP），396 - 404 FQPQNGQFI。用仅识别GP1、GP2或NP的CTL克隆与病毒感染的H-2b细胞孵育，会导致病毒变体的选择，这些变体在感染携带H-2b分子的细胞时，能逃避具有适当特异性的CTL的识别。核酸测序显示变体病毒的GP1（第38位氨基酸F→L）、GP2（第282位氨基酸G→D）或NP（第403位氨基酸F→L）发生了单个突变。当合成野生型（wt）LCMV肽和三种变体肽（GP1、GP2、NP）并进行竞争性抑制结合试验时，发现wt肽和变体肽对H-2Db的结合亲和力没有差异。用wt肽包被的未感染细胞可被适当的CTL克隆或体内致敏的大量CTL识别并裂解，但用GP1、GP2或NP变体肽包被的类似靶细胞则不能。这一结果，再结合对这些变体肽与最近解析的Db MHC I类分子三维结构的计算机图形分析，将突变残基的侧链置于MHC -肽复合物的外表面，且可被T细胞受体识别。在GP残基38或282或NP 403处进行丙氨酸替代也消除了CTL的识别和裂解作用。将任何一种突变的病毒变体接种到小鼠体内，都会产生针对其他两种未突变的GP或NP肽的有效CTL反应，这表明在体内产生具有生物学相关性的CTL逃逸病毒变体需要在不止一个且可能是所有CTL表位中选择突变，这是一个低概率事件。
