Fujieda S, Lin Y Q, Saxon A, Zhang K
Department of Medicine, UCLA School of Medicine, Los Angeles, CA 90095, USA.
J Immunol. 1996 Oct 15;157(8):3450-9.
Germ-line transcripts from Ig heavy chain loci precede the occurrence of isotype switching and are thought to play an important though still controversial role in Ig class switching. In this study, we employed a reverse transcriptase-PCR approach to detect human chimeric Ig germ-line mRNA transcripts. Multiple types of chimeric Ig germ-line transcripts (Imu-Cepsilon, Iepsilon-Cmu, Imu-Cgamma4, Igamma-Cmu, Igamma-Cepsilon, Iepsilon-Cgamma, and Igamma4-Calpha1 transcripts) were readily detected in human B cells stimulated with IL-4 alone. Sequence analysis revealed that all of these chimeric Ig germ-line transcripts represented the I exons from one Ig locus spliced to the CH exons from another locus by using consensus sequences for splicing donor and acceptor sites, indicating that they were generated through splicing machinery. In the case of stimulation of human resting B cells with IL-4 alone, the chimeric Ig germ-line transcripts are likely derived from a trans-splicing mechanism, as the extensive searching did not find evidence that Ig class-switch recombination had occurred, which alternatively could give rise to chimeric Ig mRNA by mechanisms other than trans-splicing. Similarly, an EBV-transformed gamma2 rearranged B cell line, GM1500, which produces IgG2 and contains both gamma2 productive and epsilon germ-line transcripts, also expressed chimeric germ-line RNA (Iepsilon-Cgamma2) and epsilon-productive transcripts (VDJ-Cepsilon). This line had no further sequential Sgamma2-Sepsilon rearrangements, providing evidence that the productive VDJ-Cepsilon mRNA was derived from a transcriptionally active unrearranged epsilon gene locus by trans-splicing. Taken together, these results provide possible evidence that trans-splicing of germ-line Ig RNA transcripts occurs in human B cells.
免疫球蛋白重链基因座的种系转录本先于同种型转换出现，并且被认为在免疫球蛋白类别转换中发挥着重要作用，尽管该作用仍存在争议。在本研究中，我们采用逆转录酶 - 聚合酶链反应方法来检测人嵌合免疫球蛋白种系mRNA转录本。在仅用白细胞介素 - 4刺激的人B细胞中，很容易检测到多种类型的嵌合免疫球蛋白种系转录本（Imu - Cepsilon、Iepsilon - Cmu、Imu - Cgamma4、Igamma - Cmu、Igamma - Cepsilon、Iepsilon - Cgamma和Igamma4 - Calpha1转录本）。序列分析表明，所有这些嵌合免疫球蛋白种系转录本均代表来自一个免疫球蛋白基因座的I外显子，通过使用剪接供体和受体位点的共有序列与来自另一个基因座的CH外显子拼接，这表明它们是通过剪接机制产生的。在仅用白细胞介素 - 4刺激人静止B细胞的情况下，嵌合免疫球蛋白种系转录本可能源自反式剪接机制，因为广泛的搜索未发现免疫球蛋白类别转换重组发生的证据，而类别转换重组也可通过反式剪接以外的机制产生嵌合免疫球蛋白mRNA。同样，一种EB病毒转化的γ2重排B细胞系GM1500，它产生IgG2且同时含有γ2有功能转录本和ε种系转录本，也表达嵌合种系RNA（Iepsilon - Cgamma2）和ε有功能转录本（VDJ - Cepsilon）。该细胞系没有进一步的连续Sgamma2 - Sepsilon重排，这证明有功能的VDJ - Cepsilon mRNA是通过反式剪接从转录活跃的未重排ε基因座产生的。综上所述，这些结果为种系免疫球蛋白RNA转录本在人B细胞中发生反式剪接提供了可能的证据。
