Samara Maha, Oruc Zeliha, Dougier Hei-Lanne, Essawi Tamer, Cogné Michel, Khamlichi Ahmed Amine
CNRS UMR 6101, Groupe 'Instabilité génétique et régulation transcriptionnelle', Faculté de Médecine, 2 rue du Dr Marcland, F-87025 Limoges cedex, France.
Int Immunol. 2006 Apr;18(4):581-9. doi: 10.1093/intimm/dxh400. Epub 2006 Feb 28.
Class switch recombination (CSR) is preceded by germ line transcription that initiates from promoters upstream of switch (S) sequences and terminates downstream of associated constant genes. Previous work showed that germ line transcripts and their processing are required for CSR and that germ line transcription is regulated in a major part by a regulatory region located downstream of the Ig heavy chain locus. This long-range, polarized effect can be disturbed by inserting an expressed neomycine resistance (neo(r)) gene. To contribute to a better understanding of the mechanism of such a long-distance regulation, we generated knock-in mice in which a neo(r) gene was inserted downstream of Igamma3 exon leaving intact all the necessary elements for germ line transcription and splicing. We show that the expressed neo(r) gene interferes with transcription initiation from Igamma3, and that it impairs but does not block S recombination to Cgamma3. Moreover, we show for the first time that the neo(r) gene provides through chimeric neo(r)-Cgamma3 transcripts the necessary elements for splicing of germ line transcripts by activating two novel cryptic splice sites, one in the coding region of the intronless neo(r) gene and the other in the Igamma3-Cgamma3 intron.
类别转换重组（CSR）之前会发生胚系转录，该转录从转换（S）序列上游的启动子开始，并在相关恒定基因的下游终止。先前的研究表明，胚系转录本及其加工对于CSR是必需的，并且胚系转录在很大程度上受位于免疫球蛋白重链基因座下游的调控区域调节。这种远距离的极化效应可通过插入一个表达新霉素抗性（neo(r)）基因而受到干扰。为了有助于更好地理解这种远距离调控的机制，我们构建了基因敲入小鼠，其中neo(r)基因插入到Iγ3外显子下游，同时保留了胚系转录和剪接所需的所有必要元件。我们发现，表达的neo(r)基因会干扰从Iγ3开始的转录起始，并且它会损害但不会阻断向Cγ3的S重组。此外，我们首次表明，neo(r)基因通过嵌合的neo(r)-Cγ3转录本，通过激活两个新的隐蔽剪接位点，为胚系转录本的剪接提供了必要元件，一个位于无内含子的neo(r)基因的编码区域，另一个位于Iγ3-Cγ3内含子中。
