Role of Polycomb-mediated epigenetic regulation in diffuse large B cell lymphoma

描述(由申请人提供)：越来越清楚的是，许多形式的癌症的发病机制都有显着的表观遗传学成分。许多癌症类型的特点是基因失活，基因沉默可以通过表观遗传机制，如DNA甲基化和/或获得抑制性组蛋白3修饰，如赖氨酸27或9的甲基化。H3K27的甲基化是由聚梳蛋白Ezh2催化的，多梳抑制复合体2(PRC2)的催化成分。最近，在一种特殊的癌症亚型：弥漫性大B细胞淋巴瘤生发中心B细胞亚型(GCB-DLBCL)中，检测到Ezh2催化部位的一个特定突变，该突变使其H3K27me3甲基转移酶活性过度激活，同时消除了其对H3K27单甲基化的能力。这种高度活跃的Ezh2突变在任何其他类型的淋巴瘤中都不存在，这表明它可能与GCB-DLBCL淋巴肿瘤的发生唯一相关。EZH2转录在NAOVE B细胞中被抑制，而在GC B细胞中上调。我们假设在这个阶段Ezh2的表达通常会增加，以抑制特定的基因子集，这些基因必须下调才能使NAOVE B细胞分化为GC B细胞，以及GC B细胞分化为GC B细胞后的GC B细胞。一旦Ezh2甲基化H3K27，H3K27me3就可以被多梳抑制复合体1(PRC1)识别。这种抑制性Prc1复合体的催化成分是Ring1b，它使赖氨酸119上的H2A单苷化(UH2A)，导致进一步的抑制。然而，H_2AK119也可以被其他络合物泛素化。此外，与果蝇PRC1不同，哺乳动物PRC1中的碱性4组分有许多替代的并列，导致了各种不同的PRC1复合体，这些复合体与H3K27me3具有不同的亲和力。尽管已知组蛋白结合亚基的4个哺乳动物对H3K27me3有不同的亲和力，但对具有不同组成的PRC1复合体的生物学后果知之甚少。我们假设，在GCB-DLBCL的子集中有更多的H3K27me3的直接下游结果可能是会有更多的PCR1招募和/或不同的PRC1复合体的招募。这可能会导致某些基因上更多的uH2A和/或更多的DNA甲基化。因此，这些基因很可能表现出最严重的失调。这个项目将描述在正常人类B细胞分化为GC B细胞，然后分化为GC后记忆B细胞时，Polycomb介导的表观遗传学变化。我们将进行CHIP-SEQ以确定与H3K27me3结合的PRC1组分，并将其与表达谱和DNA甲基化谱进行比较，以确定哪些基因通常以Polycomb介导的方式被抑制。我们将分析一大组GC-DLBCL，确定携带或不携带Ezh2突变的淋巴瘤的表观遗传学特征，并将其与其基因表达特征相关联。这些研究将为深入了解GCB-DLBCL中基因的失调提供新的信息，并将为在B淋巴细胞分化的后期阶段以多梳依赖的方式抑制基因的正常表观遗传机制提供新的信息。

DESCRIPTION (provided by applicant): It has become increasingly clear that the pathogenesis of many forms of cancer have a significant epigenetic component. Many cancer types are characterized by gene inactivation, and genes can be silenced through epigenetic mechanisms such as DNA methylation and/or the acquisition of repressive histone 3 modifications such as the methylation of lysines 27 or 9. Methylation of H3K27 is catalyzed by the Polycomb protein Ezh2, the catalytic component of Polycomb Repressive Complex 2 (PRC2). Recently, a specific mutation in the catalytic site of Ezh2 that hyperactivates its H3K27me3 methyltransferase activity while eliminating its ability to monomethylate H3K27 been detected with high frequency in one specific subtype of cancer: the germinal center (GC) B cell subtype of diffuse large B cell lymphoma (GCB-DLBCL). This hyperactive Ezh2 mutation is not present in any other types of lymphomas examined, suggesting that it may be uniquely relevant for GCB- DLBCL lymphomagenesis. Ezh2 transcription is repressed in naove B cells, but it is upregulated in GC B cells. We hypothesize that Ezh2 expression is normally increased at this stage to repress a specific subset of genes that must be downregulated to allow the differentiation of a naove B cell into a GC B cell, and of a GC B cell into a post-GC B cell. Once Ezh2 methylates H3K27, H3K27me3 can be recognized by the Polycomb Repressive Complex 1 (PRC1). The catalytic component of this repressive PRC1 complex is Ring1b, which monoubiquitinates H2A on lysine 119 (uH2A), leading to further repression. However, H2AK119 can be ubiquitinylated by other complexes also. Furthermore, unlike Drosophila PRC1, the basic 4 components in mammalian PRC1 have many alternative paralogs, resulting in a variety of different PRC1 complexes with different affinities for H3K27me3. Very little is known about the biological consequences of PRC1 complexes having different compositions, although it is known that the 4 mammalian paralogs of the histone binding subunit have varying affinities for H3K27me3. We hypothesize that a direct downstream consequence of more H3K27me3 in a subset of GCB-DLBCL may be that there will be more PCR1 recruitment and/or recruitment of different PRC1 complexes. This could result in more uH2A and/or more DNA methylation on some genes. Consequently, these genes are likely to display the most dysregulation. This project will characterize the Polycomb-mediated epigenetic changes that take place in normal human B cells as they differentiate into GC B cells, then into post-GC memory B cells. We will perform ChIP-seq to identify PRC1 components binding to H3K27me3, and compare this to the expression profile and DNA methylation profile to determine which genes are normally repressed in a Polycomb-mediated manner. We will analyze a large set of GC-DLBCL, will determine the epigenetic profile of lymphomas that bear or do not bear the Ezh2 mutation, and will relate this to their gene expression profile. These studies will provide insight into the dysregulation of genes in GCB- DLBCL, and will also provide novel information on the normal epigenetic mechanisms by which genes are repressed in a Polycomb-dependent manner during the late steps of B lymphocyte differentiation.