Ribosomal scanning past the primary initiation codon as a mechanism for expression of CTL epitopes encoded in alternative reading frames.
Ribosomal scanning past the primary initiation codon as a mechanism for expression of CTL epitopes encoded in alternative reading frames.
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DOI：
10.1084/jem.184.4.1319
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发表时间：
1996-10-01
期刊：
影响因子：
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通讯作者：
Eisenlohr LC
中科院分区：
文献类型：
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作者：
Bullock TN;Eisenlohr LC
越来越多的证据表明，仅限于 MHC I 类分子并被 CTL 识别的表位不需要在初级开放阅读框 (ORF) 中进行编码。此类表位已在终止密码子之后、替代阅读框 (RF) 和内含子内得到证实。我们使用一系列引入流感 A/PR/8 /34 核蛋白 (NP) 基因的移码 (FS) 来证实之前对隐性表位表达的体外观察结果，并表明它们的表达足以引发体内免疫反应。这种呈现不是由于亚显性表位、超出 FS 点的隐秘启动子的转录或内部翻译起始所致。通过向表现出最有效表达的构建体引入额外的突变，我们已经确定起始密码子通读（这里称为扫描，其中扫描核糖体绕过传统的起始密码子，在下游启动翻译）作为表位产生的可能机制。进一步的突变分析表明，虽然它应该在野生型（WT）蛋白的表达过程中起作用，但扫描并不能在我们的系统中提供处理底物的主要来源。这些发现表明（i）胸腺选择和感染过程中可用的全部自身和病原体衍生表位尚未得到充分认识；（ii）当无法识别 CTL 反应的特异性时或在传统 CTL 靶标有限的治疗情况下（可能是潜伏病毒感染和转化细胞的情况），应考虑隐性表位表达。最后，起始密码子通读为 MHC I 类分子呈递胞吐蛋白提供了合理的解释。
An increasing amount of evidence has shown that epitopes restricted to MHC class I molecules and recognized by CTL need not be encoded in a primary open reading frame (ORF). Such epitopes have been demonstrated after stop codons, in alternative reading frames (RF) and within introns. We have used a series of frameshifts (FS) introduced into the Influenza A/PR/8 /34 nucleoprotein (NP) gene to confirm the previous in vitro observations of cryptic epitope expression, and show that they are sufficiently expressed to prime immune responses in vivo. This presentation is not due to sub-dominant epitopes, transcription from cryptic promoters beyond the point of the FS, or internal initiation of translation. By introducing additional mutations to the construct exhibiting the most potent presentation, we have identified initiation codon readthrough (termed scanthrough here, where the scanning ribosome bypasses the conventional initiation codon, initiating translation further downstream) as the likely mechanism of epitope production. Further mutational analysis demonstrated that, while it should operate during the expression of wild-type (WT) protein, scanthrough does not provide a major source of processing substrate in our system. These findings suggest (i) that the full array of self- and pathogen-derived epitopes available during thymic selection and infection has not been fully appreciated and (ii) that cryptic epitope expression should be considered when the specificity of a CTL response cannot be identified or in therapeutic situations when conventional CTL targets are limited, as may be the case with latent viral infections and transformed cells. Finally, initiation codon readthrough provides a plausible explanation for the presentation of exocytic proteins by MHC class I molecules.