ACTIVATION OF THE AXL RECEPTOR TYROSINE KINASE INDUCES MITOGENESIS AND TRANSFORMATION IN 32D CELLS

axl is a transforming receptor tyrosine kinase isolated from DNA of pa tients with chronic myelogenous leukemia. Association of axl expressio n with myelogenous leukemias and its expression in primitive hematopoi etic cells suggests a role for axl in myeloid biology. To study the ce llular function of axl, we constructed a chimeric receptor tyrosine ki nase composed of the extracellular and transmembrane domains of the EG F receptor and the cytoplasmic domain of axl; this chimera was named E AK for EGFR-Axl-Kinase. The EAK chimeric receptor was expressed in the mouse myeloid progenitor cell line 32D, which is dependent on interle ukin 3 (IL-3) for proliferation and survival. Treatment of the 32D-EAK cells with EGF stimulated the tyrosine phosphorylation of the axl kin ase domain and enabled proliferation through EGF rather than IL-3. Thu s, axl can effectively couple with mitogenic signaling pathways intrin sic to 32D myeloid cells. Assay of proteins phosphorylated in response to different cytokine treatments showed that IL-3 and EGF exposure pr oduced unique profiles in the 32D-EAK cells. Furthermore, jak-2 is pho sphorylated only in response to IL-3 treatment in these cells. This su ggests that IL-3 receptor and axl transduce mitogenic signals through separate pathways. In addition, exposure of cells expressing the chime ric receptor to EGF for 19 days converted the cells to factor-independ ent growth, a phenomenon not seen with other receptor tyrosine kinases . Generation of this transformed phenotype is absolutely dependent on axl activation by foster ligand. The tyrosine phosphorylation level of the axl kinase domain in the factor-independent subclones is 40-fold greater than the factor-dependent cells. The association of a unique a xl phosphorylation level with the factor-independent phenotype suggest s that there is a threshold phosphorylation level of the axl kinase fo r transformation. The fact that activation of the axl receptor leads t o transformation of 32D cells suggests that axl can play a role in leu kemic conversion of myeloid cells, either through inappropriate expres sion or improper activation.