From the Philadelphia chromosome to the cytoskeleton: The role of BCR-ABL in modulating cell adhesion and initiating leukemia

Chronic myelogenous leukemia (CML) is a biphasic neoplasm of the bone marrow that is precipitated by the Philadelphia Chromosome, a t(9;22) balanced translocation that encodes a constitutively activated nonreceptor tyrosine kinase termed P210BCR-ABL. This oncoprotein has several intracellular functions; however, the most important effect is phosphorylation of signaling molecules through a constitutively active tyrosine kinase domain. Despite extensive knowledge of the structure and functional domains of P210 BCR-ABL, its precise role in transformation is not known. This thesis utilizes two assays to better understand the functioning of P210BCR-ABL in leukemogenesis. In the first approach, we use a murine CML model in which all wild type P210BCR-ABL recipients develop a rapidly fatal myeloproliferative disease that shares many similarities with human CML. Using this model, we have shown that the N-terminal coiled-coil domain and the grb2 binding site at Tyr177 are required to induce murine CML. A second approach utilizes a quantitative cell detachment apparatus capable of measuring small changes in cell adhesion to investigate the mechanism by which P210BCR-ABL causes abnormal cell binding. We found that P210BCR-ABL leads to increased adhesion to fibronectin in a tyrosine kinase independent manner. To better understand the role vi abnormal adhesion may play in the development of leukemia, we have attempted to correlate the effects of functional P210BCR-ABL mutants in regulating adhesion and oncogenicity. We found that localization of P210BCR-ABL to F-actin, mediated through the actin binding domain or coiled-coil domain, as well as a region between as 163 to as 210 are necessary for P210 BCR-ABL-mediated increased adhesion. We compared our adhesion results with the ability of these mutants to cause the murine CML disease and found that enhanced adhesion to fibronectin did not correlate with the ability of these mutants to cause CML. Preventing the association of P210BCR-ABL with F-actin by deleting the C-terminal actin binding domain precipitates a CML-like disease that is indistinguishable from that caused by wild-type P210BCR-ABL. Together, our results suggest that F-actin localization may play a pivotal role in modulating adhesion, but that it is dispensable for CML development.