Genetic mechanisms regulating B-lymphocyte commitment

描述(由申请人提供)：低表达的共同淋巴祖细胞的B淋巴细胞承诺的途径和分子机制在很大程度上仍不清楚。最近的研究表明，转录因子早期B细胞因子(EBF)和Pax5可能在调节B细胞谱系的启动和维持方面发挥重要作用。在没有EBF或Pax5的情况下发育的小鼠，B细胞的发育在血统承诺(EBF)之前就停止了，或者B承诺状态的维持不能持续(Pax5)。EBF可以直接结合和激活Pax5启动子的发现表明，这两个因素可能在共同的调控级联中发挥作用，控制B细胞的命运。这一观点得到了本提案中描述的研究的支持，在这些研究中，在造血干细胞中强制表达EBF或Pax5足以确定体内B细胞与T细胞的身份。所有用表达EBF或Pax5的细胞重组的动物在移植后至少10个月内，外周血和脾中90%以上的供者来源的B系细胞，基本上没有T淋巴细胞在胸腺中发生。B细胞谱系发育的强烈倾斜不是由于恶性肿瘤引起的，而是EBF或Pax5表达的特异性。这些重叠的表型进一步表明，EBF和Pax5可能在互补的途径中发挥作用，以控制B谱系确定的最早事件。为了验证这一假设，我们建议(1)定位EBF和Pax5中负责促进B细胞发育和抑制T谱系承诺的结构域，(2)在EBF-/-动物的背景下识别激活B细胞发育的EBF下游靶基因，(3)确定EBF和/或Pax5是否足以在PU缺乏的遗传背景下拯救B细胞的发育。和(4)确定通过Notch1通路的信号是否影响EBF和/或Pax5的表达或活性。这些研究将为在分子水平上控制B细胞发育的早期阶段的调节级联提供洞察力。

DESCRIPTION (provided by applicant): The pathways and molecular mechanisms responsible for B lymphocyte commitment from oligopotent common lymphoid progenitor cells remain largely unknown. Recent studies have suggested that the transcription factors, early B cell factor (EBF) and Pax5, may play a significant role in regulating both the initiation and maintenance of the B cell lineage. In mice that develop in the absence of EBF or Pax5, B cell development is arrested prior to lineage commitment (EBF) or the maintenance of the B committed state cannot be sustained (Pax5). Findings that EBF can directly bind and activate the Pax5 promoter suggest that these two factors may act in a common regulatory cascade that controls B cell fate. This notion is supported by studies described in this proposal, where enforced expression of either EBF or Pax5 in hematopoietic stem cells is sufficient to determine B versus T cell identity in vivo. All animals reconstituted with cells expressing EBF or Pax5 exhibit greater than 90 percent donor-derived B-lineage cells in the peripheral blood and spleen for at least 10 months post-transplant, with essentially no T lymphopoiesis occurring in the thymus. The strong skew in development to the B cell lineage was not due to malignancy and was specific for expression of either EBF or Pax5. The overlapping phenotypes further suggest that EBF and Pax5 may function in complementary pathways to control the earliest events in B-lineage determination. To test this hypothesis, we propose to (1) map domains within EBF and Pax5 that are responsible for both promotion of B cell development and inhibition of T-lineage commitment, (2) identify downstream target genes of EBF that activate B cell development in the context of EBF-/- animals, (3) define whether EBF and/or Pax5 are sufficient to rescue B cell development in genetic backgrounds deficient in PU. 1, Ikaros, or E2A and (4) determine whether signaling through the Notch1 pathway influences the expression or activity of EBF and/or Pax5. These studies will provide insights into the regulatory cascades that control the earliest stages of B cell development at the molecular level.