Chromatin modifications in immunoglobulin switch recombination

转换到IgE可以通过IL4和细胞表面CD40受体的联合激活来激活，IL4通过IgE开关区诱导转录，而CD40受体的激活则诱导细胞增殖。激活可以通过抗CD40抗体或CD40配体的三聚体形式产生。我们的研究主要集中在人B细胞系CL-01上，该细胞系已被报道经历了开关重组，并且最近包括了原代人B细胞。我们已经证明，IL4在CL=01细胞的IgE开关区导致转录物增加100倍，无论是未剪接的还是剪接的。我们还观察到，负责启动CSR的胞苷脱氨酶AID的转录显著增加（5倍）。

Switching to IgE can be activated by a combination of IL4, which induces transcription through the IgE switch region, and activation of the cell surface CD40 receptor, which induces cell proliferation. Activation can be produced by anti-CD40 antibody or by a trimeric form of CD40 ligand. Our studies have concentrated on the human B cell line CL-01, which has been reported to undergo switch recombination, and have more recently included primary human B cells. We have shown that IL4 causes a 100X increase in transcript, both unspliced and spliced, over the IgE switch region in CL=01 cells. We also observed a sizable (5X) increase in transcription of AID, the cytidine deaminase that is responsible for initiating CSR. A high resolution study of histone modifications in uninduced and induced CL-01 cells has been carried out, focusing on the region between the I-epsilon promoter and the 3 end of the IgE switch region. This is the most important region associated with this CSR event. The largest increases were found for the dual modification acetyl-H3K9/14 (or acetyl-H3K9 alone) and for trimethyl-H3 K4, while other modifications including histone H4 tetra-acetylation (K5/8/12/16) and H3 K27 trimethylation were not greatly altered following IL4 induction. This pattern is consistent with broad activation of the region. Furthermore, most of the modifications were unevenly distributed: for example, dimethyl-H3K4, tetraacetyl-H4, and acetyl-H3 K9/14 all were concentrated over the I-epsilon promoter. To aid in interpreting these results, the abundance of nucleosomes was mapped across the same region, and a relative depletion over I-epsilon was observed, without any further change on induction. We have shown that treatment of the cells with 5-aza-deoxycytidine, a known demethylating agent, leads to an additional increase in IL4-stimulated transcription. We have also mapped the distribution of RNA polymerase II, as well as potential sites of AID binding in this region. Working together with our collaborators in Great Britain, we have also studied histone modifications in primary human B cells from several subjects. The patterns are on average similar to that in CL-01 cells, but with notable variations between individuals, which will be the subject of future work.