Chromatin modifications in immunoglobulin switch recombination

转换为IgE可以通过IL 4的组合来激活，IL 4诱导该区域的转录，而细胞表面CD 40受体的激活诱导细胞增殖。我们的研究主要集中在人CL-01系B细胞上，据报道该细胞发生了开关重组。我们已经表明，IL 4导致在IgE转换区的转录物增加100倍，无论是未剪接的还是剪接的。我们还观察到AID（负责启动CSR的胞苷脱氨酶）的转录有相当大的（5倍）增加。 已对未诱导和诱导的CL-01细胞中的组蛋白修饰进行了高分辨率研究，重点是I-IgE启动子和IgE开关区3端之间的区域，该区域是与该CSR事件相关的最重要区域。发现乙酰基-H3 K9/14（或单独的乙酰基-H3 K9）和三甲基-H3 K4的增加最大，而其他修饰包括组蛋白H4四乙酰化（K5/8/12/16）和H3 K27三甲基化在IL 4诱导后没有显著改变。这种模式与该区域的广泛激活一致。此外，在大多数情况下，给定修饰的分布是不均匀的：例如，二甲基-H3 K4、四乙酰基-H4和乙酰基-H3 K9/14都显示出在I-CH 3启动子上的相对峰。为了解释这些结果，在同一区域绘制了核小体丰度图，并观察到相对于I-γ的消耗，诱导没有任何进一步的变化。我们现在将这些观察结果扩展到研究组蛋白变体H3.3和H2A.Z的分布。此外，我们正在研究同一区域的DNA甲基化。我们已经表明，5-氮杂-脱氧胞苷，一种已知的去甲基化剂，治疗的细胞，导致IL 4刺激的转录的额外增加。

Switching to IgE can be activated by a combination of IL4, which induces transcription in that 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 focused on the human CL-01 line of B cells, which has been reported to undergo switch recombination. We have shown that IL4 causes a 100X increase in transcript, both unspliced and spliced, over the IgE switch region. 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, the most important region associated with this CSR event. The largest increases were found for 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, the distribution of a given modification was in most cases non-uniform: for example, dimethyl-H3K4, tetraacetyl-H4, and acetyl-H3 K9/14 all showed relative peaks over the I-epsilon promoter. To interpret 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 are now extending these observations to studying the distribution of the histone variants H3.3 and H2A.Z. In addition we are studying DNA methylation in the same region. 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.