Mechanisms of human pre-B cell differentiation

 描述(由申请人提供)：通过早期B细胞分化的调节过渡对于成熟B细胞谱系的产生和选择非常重要。我们最近描述了小鼠B细胞分化早期的一个过渡期，除了BCR前和IL7R信号外，还表现出整合素依赖的基质黏附、强大的增殖能力和自我更新。转录调控因子Ikaros的缺失导致基质黏附的Pre-B细胞阶段停滞，整合素信号、基质黏附和自我更新显著增加，Pre-BCR分化信号丢失。尽管这一阶段的早期B细胞分化对包括免疫缺陷和白血病在内的人类血液系统疾病的发展也很重要，但对人类造血系统中的这一过程知之甚少，也是我们目前研究的重点。在第一个目标中，我们将评估在骨髓基质依赖系统中出现的人类B细胞前体的等级。我们将研究早期人类B细胞前体细胞在体外对增殖、扩增和分化的基质依赖性，并确定与这些事件相关的信号通路和转录因子。在免疫缺陷的NSG小鼠体内产生的人类B细胞前体的分化特性将以类似的方式进行评估。在第二个目标中，我们将研究IKAROS在人类前B细胞分化中的作用。人类B细胞前体的基因组编辑将扰乱IKZF1基因座，以模拟人类疾病的突变。在NSG小鼠体内移植后，将在体外基质培养和体内测试Ikaros突变B细胞前体细胞的分化、增殖和黏附特性。从IKAROS缺陷的人前B细胞前体和其正常对应的人产生的细胞和分子数据可以作为比较血液疾病的基线。总之，这些研究将在驱动人类发育过程及其异常表现的分子途径方面产生关键的新知识。这项工作将使我们能够在老鼠和人类之间无缝移动，以发现治疗血液疾病的更好方法。

 DESCRIPTION (provided by applicant): Regulated transitions through early B cell differentiation are of great importance for the generation and selection of the mature B cell repertoire. We have recently described a transitional phase in early mouse B cell differentiation that in addition to pre-BCR and IL7R signaling also displays integrin-dependent stromal adhesion, strong proliferative capacity and self-renewal. Loss in the transcriptional regulator Ikaros causes arrest at this stromal- adherent pre-B cell stage with a profound increase in integrin signaling, stromal adhesion and self-renewal and loss in pre-BCR differentiation signaling. Although this stage of early B cell differentiation is also important for the developmen of human hematological disorders, including immunodeficiencies and leukemias, little is known of this process in the human hematopoietic system and is the focus of our current investigation. In the first aim, we will evaluate the hierarchy of human B cell precursors that arise in a bone marrow stromal-dependent system. We will examine the in vitro stromal dependence of early human B cell precursors for proliferative expansion and differentiation and identify the signaling pathways and transcriptional factors associated with these events. The differentiation properties of human B cell precursors generated in vivo in immune-deficient NSG mice will be evaluated in a similar fashion. In the second aim, we will examine the role of IKAROS in human pre-B cell differentiation. The IKZF1 locus will be disrupted by genome editing in human B cell precursors to model mutations in human disease. IKAROS mutant B cell precursors will be tested for differentiation, proliferation and adhesion properties in in vitro stromal cultures and in vivo aftr transplantation in NSG mice. Cellular and molecular data generated from IKAROS deficient human pre-B cell precursors and its normal counterparts can serve as baselines to which hematological disorders can be compared. Together these studies will generate critical new knowledge on the molecular pathways that drive the human developmental process and its aberrant manifestations. This work will enable us to move seamlessly between the mouse and human to discover better approaches for treating hematological disorders.