Modeling ICF syndrome in mice: Role of Zbtb24 in DNA methylation and antibody production

免疫缺陷、着丝粒不稳定和面部异常（ICF）综合征是一种罕见的常染色体 隐性疾病，绝大多数病例携带DNA甲基转移酶基因突变， DNMT 3B（ICF 1）或锌指蛋白基因ZBTB 24（ICF 2）。ICF综合征的一个标志是DNA丢失 在特定基因组区域的甲基化，这被认为是潜在的其他表型的主要缺陷 异常，包括抗体缺乏症（低丙种球蛋白血症）、面部畸形和精神 迟钝ICF综合征患者通常在儿童早期死于反复感染。虽然ICF 综合征首次报道是在近四十年前，在了解 目前，研究人员对疾病的发病机制进行了研究，主要是因为缺乏适当的体外和体内模型。建模 使用Dnmt 3b突变小鼠的ICF综合征一直是一个挑战，因为Dnmt 3b的完全失活 导致胚胎死亡，携带ICF样点突变的小鼠不能重现抗体缺陷。 来自申请人实验室的初步数据显示，小鼠胚胎干细胞中的Zbtb 24消耗 （ESCs）导致ICF特征性的Dnmt 3b显著下调和DNA甲基化改变 综合征和造血谱系中条件性Zbtb 24缺失导致严重的 低丙种球蛋白血症小鼠，显然是由于缺陷的浆细胞分化或存活。的 本申请的目的是确定Zbtb 24在调节DNA甲基化中的作用， 抗体生产。中心假设是Zbtb 24通过调节Dnmt 3b表达，控制DNA 甲基化，基因表达和染色质结构，在淋巴细胞群体，是重要的 抗体生产。申请人提出使用Zbtb 24缺陷型ESC来阐明Zbtb 24缺陷型ESC的分子生物学特性。 Zbtb 24调节Dnmt 3b表达的机制，并确定Zbtb 24缺乏对Dnmt 3b表达的影响。 DNA甲基化、基因表达和染色质结构（目的1）。申请人还建议使用 Zbtb 24-突变ICF小鼠模型，以确定涉及抗体缺乏的细胞缺陷， 阐明异常DNA甲基化与基因表达、染色质结构缺陷之间的联系 以及导致抗体缺乏的分子信号传导（AIM 2）。该项目意义重大，因为 所提出的工作的结果不仅将从根本上推进对ICF的机理理解 综合征，但也可以揭示病因和缺陷的潜在其他抗体缺乏症， 包括常见变异性免疫缺陷（CVID）。这些结果也将具有普遍意义， 了解表观遗传机制在免疫和免疫疾病中的作用。该项目 潜在的翻译影响，因为拟议工作的结果可能导致识别 ICF综合征和其他免疫缺陷疾病的新治疗策略，以及创新的ICF 小鼠模型可能是有价值的临床前试验的治疗方法在未来。

The Immunodeficiency, Centromeric instability, and Facial anomalies (ICF) syndrome is a rare autosomal recessive disorder, with the vast majority of cases carrying mutations in either the DNA methyltransferase gene DNMT3B (ICF1) or the zinc finger protein gene ZBTB24 (ICF2). A hallmark of ICF syndrome is loss of DNA methylation in specific genomic regions, which is believed to be the primary defect underlying other phenotypic abnormalities, including antibody deficiency (hypogammaglobulinemia), facial dysmorphism, and mental retardation. Patients with ICF syndrome usually die of recurrent infections in early childhood. Although ICF syndrome was first reported nearly four decades ago, little progress has been made in understanding the pathogenesis of the disease, largely because of the lack of appropriate in vitro and in vivo models. Modeling ICF syndrome using Dnmt3b mutant mice has been a challenge, because complete inactivation of Dnmt3b leads to embryonic lethality and mice carrying ICF-like point mutations fail to recapitulate antibody deficiency. Preliminary data from the applicant's laboratory revealed that Zbtb24 depletion in mouse embryonic stem cells (ESCs) results in substantial Dnmt3b downregulation and DNA methylation alterations characteristic of ICF syndrome and that conditional Zbtb24 deletion in the hematopoietic lineage leads to severe hypogammaglobulinemia in mice, apparently due to defects in plasma cell differentiation or survival. The objective of this application is to determine the role of Zbtb24 in the regulation of DNA methylation and antibody production. The central hypothesis is that Zbtb24, via regulating Dnmt3b expression, controls DNA methylation, gene expression, and chromatin structure in lymphocyte populations that are important for antibody production. The applicant proposes to use Zbtb24-deficient ESCs to elucidate the molecular mechanism by which Zbtb24 regulates Dnmt3b expression and determine the impacts of Zbtb24 deficiency on DNA methylation, gene expression, and chromatin structure (aim 1). The applicant also proposes to use the Zbtb24-mutant ICF mouse model to determine the cellular defects involved in antibody deficiency and elucidate the links between aberrant DNA methylation and the defects in gene expression, chromatin structure and molecular signaling that contribute to antibody deficiency (aim 2). The project is significant, because results from the proposed work will not only fundamentally advance mechanistic understanding of ICF syndrome, but could also shed light on the etiology and defects underlying other antibody deficiency diseases, including common variable immunodeficiency (CVID). These results will also have general implications in understanding the role of epigenetic mechanisms in immunity and immunological disorders. The project has potential translational impact as well, because results from the proposed work could lead to identification of novel therapeutic strategies for ICF syndrome and other immunodeficiency diseases, and the innovative ICF mouse model could be valuable for preclinical testing of therapeutics in the future.