Conditional Mutagenesis to Study c-Myb Function

描述（申请人提供）：很明显，Myb原癌基因在造血过程中起着至关重要的作用。在每个造血谱系中，c-Myb在分化的未成熟阶段大量表达，并在分化过程中相对较晚的时间关闭。然而，对于c- Myb在造血成熟过程中所起的作用，该作用是如何介导的，以及哪些信号通路调节c- Myb的表达，几乎一无所知。在Myb位点纯合子为零的小鼠在胚胎发生的第15天死于严重的贫血。这一发现图解地证明了c-Myb在造血过程中的重要性，但排除了对分化后期c-Myb活性的研究。在造血成熟的后期阶段，缺乏一种易于处理的遗传系统允许Myb突变，这是理解c-Myb在造血过程中的作用的主要障碍。在胸腺T细胞发育过程中，CD4和CD8双阴性（DM）和双阳性（DP）胸腺细胞含有大量的c-Myb，而CD4+和CD8+单阳性（SP）细胞仅含有10 - 20%的c-Myb。在外周，c-Myb仅在静止T细胞中低水平表达，但在细胞周期的G1晚期/ SD早期增殖T细胞中表达增加。为了开始了解c- Myb在T细胞发育过程中所起的作用，我们生产了携带Myb等位基因的小鼠，该等位基因被Cre重组酶靶向loxP位点进行删除。通过将这些小鼠培育成可获得的小鼠品系，将Cre的表达引导到胸腺中T细胞发育的不同阶段，我们确定了T细胞发育过程中需要c-Myb的临界点：从DN到DP发育阶段的过渡，为了预先选择DP细胞的存活和CD4 SP胸腺细胞的分化。最近，我们已经确定了c-Myb在维持T细胞稳态和介导T细胞辅助功能中的作用。本研究的目的是了解c-Myb在T细胞发育DN阶段、外周T稳态和辅助T细胞功能中的功能基础。

DESCRIPTION (provided by applicant): It is clear that the Myb protooncogene plays a crucial role during hematopoiesis. In each hematopoietic lineage, c-Myb is abundantly expressed at the immature stages of differentiation and is turned off at a relatively late time during the differentiation process. However, virtually nothing is known about what role c- Myb plays during hematopoietic maturation, how that role is mediated or what signaling pathways regulate c- Myb expression. Mice that are homozygous null at the Myb locus die at day fifteen during embryogenesis from a severe anemia. This finding graphically demonstrated the significance of c-Myb during hematopoiesis but has precluded study of c-Myb activity at the later stages of differentiation. The lack of a tractable genetic system that will allow mutation of Myb during the later stages of hematopoietic maturation has been a major impediment to understanding the role of c-Myb during hematopoiesis. During T cell development in the thymus, CD4 and CD8 double negative (DM) and double positive (DP) thymocytes contain abundant amounts of c-Myb while CD4+ and CD8+ single positive (SP) cells contain only 10 to 20% as much c-Myb. In the periphery, c-Myb is expressed at only low levels in resting T cells, but expression increases in proliferating T cells in late G1/early SD phase of the cell cycle. To begin to understand the role played by c- Myb during T cell development we have produced mice that carry a Myb allele targeted with loxP sites for deletion by the Cre recombinase. By breeding these mice to available mouse strains that direct Cre expression to different stages of T cell development in the thymus we have defined critical points during T cell development where c-Myb is required: transition from the DN to the DP stage of development, for the survival of pre-selection DP cells and the differentiation of CD4 SP thymocytes. More recently we have identified roles for c-Myb in maintaining T cell homeostasis and mediating T cell helper function. The goals of this proposal are to understand the basis for c-Myb function during the DN stage of T cell development, in peripheral T homeostasis and helper T cell function.