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: 7967408
• 负责人: MARTIN F. GELLERT
• 金额: $ 21.51万
• 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/7967408
• 关键词: Acetylation; Allergic; Antibodies; B-Lymphocytes; Binding; CD40 Ligand; Cell Line; Cell Proliferation; Cell surface; Cells; Cytidine Deaminase; DNA Methylation; Deoxycytidine; Event; Genetic Transcription; Histone H4; Histones; Human; IL4 gene; IgE; Immunoglobulin Class Switching; Immunoglobulin Switch Recombination; Immunoglobulins; Maps; Mediating; Modification; Nucleosomes; Pattern; RNA Polymerase II; RNA Splicing; Relative (related person); Reporting; Resolution; Site; TNFRSF5 gene; Transcript; Variant; Work; chromatin modification; constant region gene; cytokine; histone modification; promoter; receptor; response

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 concentrate on the human B cell line CL-01, 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. 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 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. Further work is in progress to determine potential sites of AID binding in this region, as well as the distribution of RNA polymerase II. Efforts are also underway to find conditions that stimulate switching to IgE.

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。活化可以通过抗CD 40抗体或通过三聚体形式的CD 40配体产生。我们的研究集中在人B细胞系CL-01，这已被报道经历开关重组。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。 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。这是与此次CSR活动相关的最重要区域。发现双重修饰乙酰基-H3 K9/14（或单独的乙酰基-H3 K9）和三甲基-H3 K4的最大增加，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，大多数修饰是不均匀分布的：例如，二甲基-H3 K4、四乙酰基-H4和乙酰基-H3 K9/14都集中在I-STAT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丰度图，并观察到相对于I-γ的消耗，诱导时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。进一步的工作正在进行中，以确定该区域中AID结合的潜在位点，以及RNA聚合酶II的分布。 也正在努力寻找刺激转换为IgE的条件。