Functions and mechanisms of transcriptional coactivator OCA-B in B cell development and lymphomagenesis

项目摘要 功能性免疫系统是人类健康的核心，同时，正常淋巴细胞的改变也是人类健康的核心。 发育程序可能导致免疫缺陷、自身免疫性疾病和各种淋巴瘤。 生发中心（GC）是在抗原诱导过程中在次级淋巴器官中形成的显微解剖结构。 刺激免疫反应。GC是B细胞通过克隆扩增、体细胞增殖和增殖而成熟的场所。 超突变和亲和力介导选择-因此在抗体的分化中起关键作用 分泌细胞和记忆B细胞。OCA-B是一种独特的转录辅激活因子，在胃癌中高度表达 B细胞和GC衍生的淋巴瘤，如弥漫性大B细胞淋巴瘤和伯基特淋巴瘤。体内 研究表明OCA-B对于抗原依赖性B细胞分化（包括 GC形成）和次级免疫球蛋白基因的正常表达。然而，人们对此知之甚少。 OCA-B在正常GC形成和GC衍生淋巴瘤进展中的确切作用。此外，该机制 OCA-B通过什么途径调节其靶基因也不清楚。为了解决这些问题，我们的初步研究 已经证明了（i）OCA-B在远端增强子上的功能占据，特别是大基因座对照 区域（LCR）的BCL 6基因编码的GC形成的主调节器;（ii）直接相互作用的OCA- B与介体共激活因子复合物的MED 1亚基和GC特异性转录因子（MEF 2 B） 对于BCL 6表达是重要的，和（iii）体内GC B细胞分化的B细胞内在Oca-B依赖性。 根据我们以前的OCA-B研究和最近的初步数据，我们假设， OCA-B在GC形成中的作用涉及通过募集和激活BCL 6通过其远端增强子区域， 浓缩转录因子和辅因子，包括染色质/表观遗传因子，其促进增强子- 启动子相互作用我们还假设OCA-B在GC衍生的淋巴瘤发生中起关键作用。到 为了解决这些问题，我们将研究（i）OCA-B的功能和作用机制，通过相互作用 与MED 1/Mediator和MEF 2 B一起，在离体GC B和淋巴瘤细胞中对BCL 6 LCR/超级增强子的作用， (ii)OCA-B依赖的转录共激活通过严格的生物化学作用机制， 体外转录测定;（iii）OCA-B在体内GC B细胞分化中的作用机制，包括 关键靶基因和调控网络的鉴定;（iv）OCA-B在B细胞淋巴瘤进展中的作用 in vivo.我们将使用互补的生物化学，基因组学，生物信息学，遗传学，基因编辑，基于细胞和 体内（小鼠模型）方法。这些目标的完成将促进我们对分子生物学的理解。 OCA-B在GC特异性转录调节、GC B细胞分化和GC-1中的作用机制 衍生的B细胞淋巴瘤形成。值得注意的是，它还将为治疗B的新治疗策略提供线索 细胞淋巴瘤

PROJECT SUMMARY A functional immune system is central to human health and, at the same time, alterations of normal lymphoid developmental programs can lead to immune deficiencies, autoimmune diseases and a variety of lymphomas. Germinal centers (GCs) are microanatomical structures formed in secondary lymphoid organs during antigen- stimulated immune responses. GCs are the sites of B cell maturation through clonal expansion, somatic hypermutation, and affinity-mediated selection -- and therefore play a critical role in differentiation of antibody secreting cells and memory B cells. OCA-B is a unique transcriptional coactivator that is highly expressed in GC B cells and GC-derived lymphomas such as diffuse large B cell lymphoma and Burkitt’s lymphoma. In vivo studies have demonstrated that OCA-B is essential for both antigen-dependent B cell differentiation (including GC formation) and normal expression of secondary immunoglobulin genes. However, little is known about the exact role of OCA-B in normal GC formation and GC-derived lymphoma progression. In addition, the mechanism by which OCA-B regulates its target genes is also unclear. In addressing these issues, our preliminary studies have demonstrated (i) a functional occupancy of OCA-B on distal enhancers, in particular the large locus control region (LCR) of the BCL6 gene that encodes a master regulator of GC formation; (ii) direct interactions of OCA- B with the MED1 subunit of the Mediator coactivator complex and with a GC-specific transcription factor (MEF2B) important for BCL6 expression and (iii) a B cell-intrinsic Oca-B dependency for GC B cell differentiation in vivo. Based on our previous OCA-B studies and recent preliminary data, we hypothesize that a predominant role of OCA-B in GC formation involves activation of BCL6 through its distal enhancer region by recruiting and concentrating transcription factors and cofactors, including chromatin/epigenetic factors, that facilitate enhancer- promoter interactions. We also hypothesis that OCA-B plays a critical role in GC-derived lymphomagenesis. To address these issues, we will investigate (i) the function and mechanism of action of OCA-B, through interactions with MED1/Mediator and MEF2B, on the BCL6 LCR/super-enhancer in both ex vivo GC B and lymphoma cells, (ii) the mechanism of action of OCA-B-dependent transcriptional coactivation through rigorous biochemical in vitro transcription assays; (iii) the mechanism of action of OCA-B in GC B cell differentiation in vivo, including identification of key target genes and regulatory networks; (iv) the role of OCA-B in B cell lymphoma progression in vivo. We will use complementary biochemical, genomic, bioinformatic, genetic, gene-editing, cell-based and in vivo (mouse model) approaches. Completion of these aims will advance our understanding of the molecular mechanism of action of OCA-B in GC-specific transcriptional regulation, GC B cell differentiation, and GC- derived B cell lymphomagenesis. Notably, it also will provide clues to new therapeutic strategies for treating B cell lymphomas.