Chromatin modifications in immunoglobulin switch recombination

免疫球蛋白开关重组中的染色质修饰

• 批准号: 8148771
• 负责人: MARTIN F. GELLERT
• 金额: $ 44.58万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8148771
• 关键词: Immunoglobulin Switch Recombination; chromatin modification

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.

切换到IgE可以通过IL4的组合来激活，IL4的组合通过IgE开关区域诱导转录，并激活细胞表面CD40受体，从而诱导细胞增殖。激活可以由抗CD40抗体或CD40配体三聚体形式产生。我们的研究集中在人类B细胞系CL-01上，据报道该细胞系进行了转换重组，并且最近包括原代人B细胞。我们已经表明，IL4在Cl = 01个细胞中的IgE开关区域上引起的转录本（未贴合和剪接）增加了100倍。我们还观察到了负责启动CSR的胞苷脱氨酶的AID转录的大幅度（5倍）。 已经进行了对未诱导和诱导的CL-01细胞中组蛋白修饰的高分辨率研究，重点是I-EPSILON启动子和IgE开关区域的3端之间的区域。这是与此CSR事件相关的最重要区域。发现双重修饰的乙酰基-H3K9/14（或单独的乙酰H3K9）和三甲基-H3 K4的最大增加，而包括组蛋白H4二乙酰化（K5/8/12/16）（K5/8/12/16）和H3 K27甲基化（H3 K27甲基化）在IL4诱导后并没有发生很大的改变。这种模式与该区域的广泛激活一致。此外，大多数修饰分布不均：例如，二甲基-H3K4，四乙酰基-H4和乙酰基-H3 K9/14都集中在I-EPSILON启动子上。为了帮助解释这些结果，在同一区域绘制了核小体的丰度，并且观察到I-Epsilon的相对耗竭，而没有进一步改变诱导。我们已经表明，用5-aza-脱氧胞苷（一种已知的脱甲基剂）治疗细胞会导致IL4刺激的转录增加。我们还绘制了RNA聚合酶II的分布以及该区域中辅助结合的潜在位点。 与我们在英国的合作者合作，我们还研究了来自多个受试者的原代人B细胞中的组蛋白修饰。这些模式平均与CL-01细胞中的模式相似，但是个体之间有明显的差异，这将是未来工作的主题。