JAK2 MEDIATED SIGNAL TRANSDUCTION

这项研究计划的长期目标是表征酪氨酸激酶Jak2下游的信号通路，这对造血细胞的发育至关重要。Jak2在造血中起着关键作用，在缺乏功能性Jak2基因的小鼠中，几个髓系的完全缺失证明了这一点。此外，它在血液学发展中的重要性被最近的发现所强调，即几种不同血液学恶性肿瘤患者的白血病细胞含有一种由转录因子TEL融合到JAK2催化结构域的异常融合蛋白。虽然Jak2激活的信号级联尚未被很好地表征，但最近的数据表明Jak2的酪氨酸自磷酸化在功能上很重要。这个提议的一个关键目标是分析JAK2的自磷酸化位点。这将在一定程度上通过Jak2野生型或酪氨酸-苯丙氨酸点突变体重构Jak2-/-胎儿肝细胞来实现，并通过体外集落形成或体内重构能力对细胞进行功能分析。另一个重要的目标将是识别和表征直接与磷酸化Jak2相互作用的分子。这些分子是Jak2效应分子的良好候选分子。因此，本研究从两个方面着手研究Jak2信号的问题：首先通过分析Jak2本身，其次通过分析与Jak2相互作用的分子。

The long term objectives of this research proposal are to characterize the signaling pathways downstream of the tyrosine kinase Jak2 which are critical for hematopoietic cell development. That Jak2 plays a critical role in hematopoiesis is demonstrated by the complete absence of several myeloid lineages in mice lacking a functional JAK2 gene. Furthermore, its importance in hematological development is underscored by the recent discovery that leukemic cells of several patients with different hematological malignancies contain an abnormal fusion protein comprised of the transcription factor TEL fused to the catalytic domain of JAK2. Although the signaling cascades activated by Jak2 are not well characterized, recent data suggests that tyrosine autophosphorylation of Jak2 is functionally important. One critical goal of this proposal is to analyze the autophosphorylation sites of JAK2. This will be accomplished, in part, by reconstituting Jak2-/- fetal liver cells with either wild-type or tyrosine-to-phenylalanine point mutants of Jak2, and functionally analyzing the cells by in vitro colony formation or in vivo reconstitution ability. Another important goal will be to identify and characterize molecules which interact directly with phosphorylated Jak2. Such molecules are good candidate Jak2 effector molecules. Thus, this study approaches the problem of studying Jak2 signaling on two fronts: firstly by analyzing Jak2 itself, and secondly, by analyzing the molecules which interact with Jak2.