Control of B cell Development by YY1

描述（由申请人提供）：阴阳-1 （YY1）是一种多功能转录因子，其名称来源于其根据DNA结合位点背景或细胞类型抑制或激活转录的能力。YY1对B细胞发育很重要，因为B细胞谱系中YY1的条件敲除会导致早期B细胞缺陷，很少有成熟的B细胞。初始阻滞发生在前b细胞阶段，与免疫球蛋白（Ig）位点缺失有关，大规模染色体收缩导致远端可变(V)区基因重排减少。然而，其他B细胞缺陷也很明显，因为重排的Ig基因只能部分恢复突变表型。我们鉴定YY1为哺乳动物Polycomb Group （PcG）蛋白，为这些作用的机制基础提供了潜在的见解。有趣的是，YY1条件敲除小鼠显示出与敲除PcG蛋白EZH2相似的B细胞缺陷表型。我们开发了一个YY1突变体（YY1 REPO），该突变体特异性地削弱YY1 PcG功能，并发现，与野生型YY1相比，YY1 REPO突变体在YY1缺失的骨髓重建系统中有效地挽救了B细胞的发育，而YY1 REPO突变体无法支持有效的B细胞发育。我们还发现YY1的REPO结构域是YY1 PcG功能所必需的，能够与参与染色体收缩的凝缩蛋白和粘聚蛋白复合物相互作用。基于这些发现和各种其他数据，我们假设YY1将PcG和凝缩蛋白募集到免疫球蛋白位点，导致染色体收缩，这是远端可变区基因重排所必需的。与这一假设一致，我们发现YY1可以结合Ig kappa基因座内的多个位点，并且值得注意的是，EZH2和凝缩蛋白与YY1在这些位点共定位。凝缩蛋白亚基SMC4的敲除导致Ig重排减少，这表明这些相互作用与Ig基因座收缩直接相关。我们将利用YY1 REPO突变体结合多种新的YY1-null条件敲除系统来评估YY1 PcG功能在体内B细胞发育中的作用（目的1）。我们还将使用我们的YY1-null重构系统来确定YY1 PcG功能在Ig位点收缩和远端V区基因重排中的作用（目的2）。最后，我们将探讨YY1在免疫球蛋白位点收缩中的作用机制（目的3）。这些研究将为YY1 PcG功能在B细胞发育和免疫功能关键过程中的作用提供关键的机制细节。了解YY1的这些功能可能为PcG功能障碍或异常干细胞自我更新引起的疾病提供多种可能的治疗干预途径。

DESCRIPTION (provided by applicant): Yin Yang-1 (YY1) is a multifunctional transcription factor that derives it name from its ability to either repress or activate transcription depending upon DNA binding site context or cell type. YY1 is important for B cell development, as conditional knock-out of YY1 in the B cell lineage results in an early B cell defect with very few mature B cells. The initial block is at the pro-B cell stage and is associated with absence of immunoglobulin (Ig) locus large-scale chromosomal contraction resulting in a reduction in rearrangement of distal variable (V) region genes. However, other B cell defects are also evident because a rearranged Ig gene only partially reverts the mutant phenotype. Potential insight into the mechanistic basis for these effects is provided by our identification of YY1 as a mammalian Polycomb Group (PcG) protein. Interestingly, YY1 conditional knockout mice show a B cell defect phenotype similar to that seen by knock-out of the PcG protein, EZH2. We developed a YY1 mutant (YY1 REPO) that specifically ablates YY1 PcG function, and found that, in contrast to wild-type YY1 that efficiently rescued B cell development in a YY1-null bone marrow reconstitution system, the YY1 REPO mutant is incapable of supporting efficient B cell development. We also found that the YY1 REPO domain, necessary for YY1 PcG function, is able to physically interact with condensin and cohesin complex proteins involved in chromosomal contraction. Based upon these findings, and a variety of additional data, we hypothesize that YY1 recruits PcG and condensin proteins to the immunoglobulin loci resulting in the chromosomal contraction needed for rearrangement of distal variable region genes. Consistent with this hypothesis, we found that YY1 can bind to multiple sites within the Ig kappa locus and that, remarkably, EZH2 and condensin proteins colocalize with YY1 at these sites. Knock-down of condensin subunit SMC4 resulted in reduced Ig rearrangement suggesting that these interactions are directly related to Ig locus contraction. We will utilize the YY1 REPO mutant in conjunction with multiple novel YY1-null conditional knock-out systems to assess the role of YY1 PcG function in B cell development in vivo (Aim 1). We will also use our YY1-null reconstitution systems to determine the role of YY1 PcG function in Ig locus contraction and rearrangement of distal V region genes (Aim 2). Finally, we will explore the mechanism of YY1 function in immunoglobulin locus contraction (Aim 3). These studies will provide key mechanistic detail on the role of YY1 PcG function for B cell development and for processes critical for immune function. Understanding these functions of YY1 may provide multiple avenues for possible therapeutic intervention in diseases caused by PcG dysfunction, or aberrant stem cell self renewal. PUBLIC HEALTH RELEVANCE: Our studies will determine the mechanism of YY1 function needed for hematopoietic stem cell development, B cell development, and for Ig locus contraction. These studies may provide new mechanisms for enhancing the success of bone marrow transplantation treatments, and may provide new targets for diseases caused by PcG proteins or stem cell dysfunction.