aAIDS Institute, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong bInstitutes of Biomedical Sciences, Shanghai Public Health Clinical Center, Fudan University, Shanghai cDivision of Research on Virology and Immunology, National Center for AIDS/STD Control and Prevention, China CDC, Beijing, China.
AIDS. 2013 Oct 23;27(16):2529-39. doi: 10.1097/QAD.0000000000000015.
Developing an effective HIV-1 vaccine that elicits broadly neutralizing HIV-1 human antibodies (bnAbs) remains a challenging goal. Extensive studies on HIV-1 have revealed various strategies employed by the virus to escape host immune surveillance. Here, we investigated the human antibody gene repertoires of uninfected and HIV-1-infected individuals at genomic DNA (gDNA) and cDNA levels by deep sequencing followed by high-throughput sequence analysis to determine the frequencies of putative germline antibody genes of known HIV-1 monoclonal bnAbs (bnmAbs).
Combinatorial gDNA and cDNA antibody libraries were constructed using the gDNAs and mRNAs isolated from uninfected and HIV-1-infected human peripheral blood mononuclear cells (PBMCs). All libraries were deep sequenced and sequences analysed using IMGT/HighV-QUEST software (http://imgt.org/HighV-QUEST/index.action). The frequencies of putative germline antibodies of known bnmAbs in the gDNA and cDNA libraries were determined.
The human gDNA antibody libraries were more diverse in heavy and light chain V-gene lineage usage than the cDNA libraries, indicating that the human gDNA antibody gene repertoires may have more potential than the cDNA repertoires to develop HIV-1 bnAbs. The frequencies of the heavy and kappa and lambda light chain variable regions with identical V(D)J recombinations to known HIV-1 bnmAbs were extremely low in human antibody gene repertoires. However, we found relatively high frequencies of the heavy and kappa and lambda light chain variable regions that used the same V-genes and had the same CDR3 lengths as known HIV-1 bnmAbs regardless of (D)J-gene usage. B-cells bearing B-cell receptors of such heavy and kappa and lambda light chain variable regions may be stimulated to induce HIV-1 bnAbs.
开发一种能诱导广泛中和 HIV-1 人类抗体（bnAbs）的有效 HIV-1 疫苗仍然是一个具有挑战性的目标。对 HIV-1 的广泛研究揭示了病毒逃避宿主免疫监视的各种策略。在这里，我们通过深度测序和高通量序列分析，在基因组 DNA（gDNA）和 cDNA 水平上研究了未感染和 HIV-1 感染个体的人类抗体基因库，以确定已知 HIV-1 单克隆 bnAbs（bnmAbs）的潜在 germline 抗体基因的频率。
使用从未感染和 HIV-1 感染的人类外周血单核细胞（PBMC）中分离的 gDNA 和 mRNA 构建组合 gDNA 和 cDNA 抗体文库。所有文库均进行深度测序，并使用 IMGT/HighV-QUEST 软件（http://imgt.org/HighV-QUEST/index.action）进行序列分析。确定 gDNA 和 cDNA 文库中已知 bnmAbs 的潜在 germline 抗体的频率。
与 cDNA 文库相比，人类 gDNA 抗体文库在重链和轻链 V 基因谱系的使用上更加多样化，这表明人类 gDNA 抗体基因库可能比 cDNA 库更有潜力开发 HIV-1 bnAbs。已知 HIV-1 bnmAbs 的重链和 kappa 和 lambda 轻链可变区具有相同 V(D)J 重组的频率在人类抗体基因库中极低。然而，我们发现相对较高频率的重链和 kappa 和 lambda 轻链可变区使用相同的 V 基因并且具有相同的 CDR3 长度作为已知的 HIV-1 bnmAbs，而不管（D）J 基因的使用情况如何。携带这种重链和 kappa 和 lambda 轻链可变区的 B 细胞受体的 B 细胞可能会被刺激以诱导 HIV-1 bnAbs。
