Mina and Everard Goodman Faculty of Life Sciences, Bar Ilan University, Ramat Gan 52900, Israel.
J Immunol. 2013 Jun 1;190(11):5567-77. doi: 10.4049/jimmunol.1201929. Epub 2013 Apr 29.
The Ab repertoire is not uniform. Some variable, diversity, and joining genes are used more frequently than others. Nonuniform usage can result from the rearrangement process, or from selection. To study how the Ab repertoire is selected, we analyzed one part of diversity generation that cannot be driven by the rearrangement mechanism: the reading frame usage of DH genes. We have used two high-throughput sequencing methodologies, multiple subjects and advanced algorithms to measure the DH reading frame usage in the human Ab repertoire. In most DH genes, a single reading frame is used predominantly, and inverted reading frames are practically never observed. The choice of a single DH reading frame is not limited to a single position of the DH gene. Rather, each DH gene participates in rearrangements of differing CDR3 lengths, restricted to multiples of three. In nonproductive rearrangements, there is practically no reading frame bias, but there is still a striking absence of inversions. Biases in DH reading frame usage are more pronounced, but also exhibit greater interindividual variation, in IgG(+) and IgA(+) than in IgM(+) B cells. These results suggest that there are two developmental checkpoints of DH reading frame selection. The first occurs during VDJ recombination, when inverted DH genes are usually avoided. The second checkpoint occurs after rearrangement, once the BCR is expressed. The second checkpoint implies that DH reading frames are subjected to differential selection. Following these checkpoints, clonal selection induces a host-specific DH reading frame usage bias.
抗体的可变区（Ab） repertoire 并不统一。一些可变区、多样性和连接基因比其他基因使用得更频繁。非均匀使用可能是由于重排过程或选择造成的。为了研究 Ab repertoire 是如何被选择的，我们分析了多样性产生的一个不能由重排机制驱动的部分：DH 基因的阅读框使用。我们使用了两种高通量测序方法，多个对象和先进的算法来测量人类 Ab repertoire 中的 DH 阅读框使用情况。在大多数 DH 基因中，主要使用单一阅读框，几乎从未观察到倒置阅读框。选择单一的 DH 阅读框不受 DH 基因单一位置的限制。相反，每个 DH 基因都参与不同 CDR3 长度的重排，限制在三个的倍数。在非生产性重排中，实际上没有阅读框偏向，但仍然存在倒置的明显缺失。DH 阅读框使用的偏向在 IgG(+)和 IgA(+) B 细胞中比在 IgM(+) B 细胞中更为明显，但也表现出更大的个体间变异性。这些结果表明，DH 阅读框选择有两个发育检查点。第一个发生在 VDJ 重组期间，此时通常避免倒置的 DH 基因。第二个检查点发生在重排后，当 BCR 表达时。第二个检查点意味着 DH 阅读框受到差异选择。在这些检查点之后，克隆选择诱导宿主特异性的 DH 阅读框使用偏向。
