Centro Nacional de Microbiología, Instituto de Salud Carlos III, 28220 Majadahonda, Spain.
Proteomics Unit, Spanish National Biotechnology Center (CNB-CSIC), 28049 Madrid, Spain.
Int J Mol Sci. 2021 Sep 29;22(19):10503. doi: 10.3390/ijms221910503.
Identification of a natural human leukocyte antigen (HLA) ligandome is a key element to understand the cellular immune response. Advanced high throughput mass spectrometry analyses identify a relevant, but not complete, fraction of the many tens of thousands of self-peptides generated by antigen processing in live cells. In infected cells, in addition to this complex HLA ligandome, a minority of peptides from degradation of the few proteins encoded by the viral genome are also bound to HLA class I molecules. In this study, the standard immunopeptidomics strategy was modified to include the classical acid stripping treatment after virus infection to enrich the HLA ligandome in virus ligands. Complexes of HLA-B27:05-bound peptide pools were isolated from vaccinia virus (VACV)-infected cells treated with acid stripping after virus infection. The HLA class I ligandome was identified using high throughput mass spectrometry analyses, yielding 37 and 51 natural peptides processed and presented untreated and after acid stripping treatment VACV-infected human cells, respectively. Most of these virus ligands were identified in both conditions, but exclusive VACV ligands detected by mass spectrometry detected on acid stripping treatment doubled the number of those identified in the untreated VACV-infected condition. Theoretical binding affinity prediction of the VACV HLA-B27:05 ligands and acute antiviral T cell response characterization in the HLA transgenic mice model showed no differences between HLA ligands identified under the two conditions: untreated and under acid stripping condition. These findings indicated that acid stripping treatment could be useful to identify HLA class I ligands from virus-infected cells.
鉴定天然人类白细胞抗原 (HLA) 配体组是理解细胞免疫反应的关键要素。先进的高通量质谱分析可以识别出由活细胞中抗原加工产生的成千上万种自身肽中的一个相关但不完整的部分。在受感染的细胞中，除了这种复杂的 HLA 配体组外，少数来自病毒基因组编码的少数几种蛋白质降解的肽也与 HLA Ⅰ类分子结合。在这项研究中，对标准免疫肽组学策略进行了修改，包括在病毒感染后进行经典的酸处理，以富集 HLA 配体组中的病毒配体。用酸处理后从痘苗病毒 (VACV) 感染的细胞中分离出与 HLA-B27:05 结合的肽池复合物。使用高通量质谱分析鉴定 HLA Ⅰ类配体组，分别从未经酸处理和酸处理 VACV 感染细胞中鉴定出 37 个和 51 个天然肽。这些病毒配体大多数在两种条件下都被鉴定出来，但通过质谱检测到的仅在酸处理条件下检测到的 VACV 特异性配体数量是未经酸处理 VACV 感染条件下鉴定出的配体数量的两倍。对 VACV HLA-B27:05 配体的理论结合亲和力预测和 HLA 转基因小鼠模型中的急性抗病毒 T 细胞反应特征表明，在未经处理和酸处理条件下鉴定出的 HLA 配体之间没有差异。这些发现表明酸处理可以用于鉴定病毒感染细胞中的 HLA Ⅰ类配体。
