SIGNAL TRANSDUCTION PATHWAYS IN STABLE PHASE CHRONIC MYELOID LEUKEMIA CELLS

费城染色体易位产生一个嵌合癌基因， 其中BCR基因和c-ABL基因融合。 这个产品 癌基因p210 BCR/ABL具有升高的ABL酪氨酸激酶活性， 并磷酸化几种细胞蛋白质。 BCR/ABL 转化造血细胞，诱导因子独立性，减少 凋亡，并改变CML细胞的粘附。 然而，在一个生化 BCR/ABL转化骨髓细胞的机制很差， 明白 已经鉴定了BCR/ABL激酶的几种底物， 包括c-BCR、p120rasGAP、c-CBL、p52SHC、p93FES、p95VAV、p125FAK、 p68桩蛋白和p72 SHPTP 2。 此外，BCR/ABL已被证明直接结合 在BCR的Y177处与GRB 2结合，因此可能激活p21 ras。 然而，很难确定其中任何一个的重要性。 这些潜在的BCR/ABL底物，部分原因是 研究一种具有许多潜在信号基序的大蛋白质。 一 简化BCR/ABL生物学的方法是检查初级人类 CML细胞，而不是过度表达BCR/ABL的细胞系。 有趣的是，在原代白血病细胞中， 这表明，在原发性肝癌中的研究， CML细胞，而不是组织培养细胞系，可能更可靠， 识别重要信号通路的术语。 在初步研究中 我们发现只有一个酪氨酸磷蛋白与 CML中性粒细胞中的BCR/ABL，最近被确定为CRKL，AN sh2/sh3 "衔接"蛋白。 CRKL通过其SH3结构域与BCR/ABL结合。 在 在进一步的研究中，我们发现了两种细胞蛋白， CML细胞中CRKL S2结构域。 第一种蛋白质是局灶性的 第二种是称为CAS的130 kDa蛋白， CRK相关底物 在初步研究中，我们克隆了 和鸡pavilion基因，并确定了结合CRKL SH2和 其他蛋白质。 这里要检验的中心假设是， BCR/ABL与CRKL衔接蛋白的相互作用在 稳定期CML的发病机制。 我们的初步数据表明，CRKL， 通过与桩蛋白和/或CAS结合，可能激活一种途径， 调节整合素功能、活力或增殖，并且这些 将对假设进行检验。 预计这些结果将 提高我们对CML发病机制的认识。

The Philadelphia chromosome translocation generates a chimeric oncogene in which the BCR gene and the c-ABL genes are fused. The product of this oncogene, p210BCR/ABL, has elevated ABL tyrosine kinase activity, relocates to the cytoskeleton, and phosphorylates several cellular proteins. BCR/ABL transforms hematopoietic cells, induces factor-independence, reduction of apoptosis, and alters adhesion of CML cells. However, at a biochemical level, the mechanisms by which BCR/ABL transforms myeloid cells are poorly understood. Several substrates of the BCR/ABL kinase have been identified, including c-BCR, p120rasGAP, c-CBL, p52SHC, p93FES, p95VAV, p125FAK, p68paxillin, and p72SHPTP2. Also, BCR/ABL has been shown to bind directly to GRB2 at Y177 of BCR, and therefore potentially activating p21 ras. However, it has been difficult to determine the significance of any of these potential BCR/ABL substrates, in part due to the complexity of studying a large protein with many potential signaling motifs. One approach to simplifying BCR/ABL biology has been to examine primary human CML cells, rather than cell lines made to overexpress BCR/ABL. Interestingly, in primary leukemic cells, there are only a few proteins which are phosphorylated by BCR/ABL. This suggests that studies in primary CML cells, rather than tissue culture cell lines, may be more reliable in terms of identifying important signaling pathways. In preliminary studies we found that there is only a single tyrosine phosphoprotein complexed with BCR/ABL in CML neutrophils, recently identified as CRKL, AN sh2/sh3 "adapter" protein. CRKL binds to BCR/ABL through its SH3 domain. In additional studies, we have identified two cellular proteins which bind to the CRKL S2 domain in CML cells. The first protein is a component of focal adhesions, p68 paxillin, and the second is a 130 kDa protein termed CAS for "CRK associated substrate". In preliminary studies, we cloned the human and chicken pavilion genes, and identified sites for binding CRKL SH2 and other proteins. The central hypothesis to be tested here is that the interaction of BCR/ABL with the CRKL adapter protein is important in the pathogenesis of stable phase CML. Our preliminary data suggest that CRKL, through binding to paxillin and or CAS, may be activating a pathway which regulates integrin function, viability, or proliferation, and these hypotheses will be tested. It is anticipated that these results will improve our understanding of the pathogenesis of CML.