CONSTITUTIVELY ACTIVATING MUTATIONS OF C-KIT RECEPTOR TYROSINE KINASECONFER FACTOR-INDEPENDENT GROWTH AND TUMORIGENICITY OF FACTOR-DEPENDENT HEMATOPOIETIC-CELL LINES

The c-kit receptor tyrosine kinase (KIT) is activated upon ligand bind ing, thereby leading to a variety of signaling events that play a fund amental role in hematopoiesis. In addition to ligand-dependent activat ion, we have previously shown that KIT is constitutively activated in a ligand-independent manner by two point mutations, Vat-559 --> Gly (G 559) mutation in the juxtamembrane domain and Asp-814 --> Val (V814) m utation in the phosphotransferase domain. To investigate the biochemic al consequence and biologic significance of these mutations, retrovira l vectors encoding KITG559 Or KITV814 were introduced into murine pro- B-type Ba/F3 cells and myeloid FDC-P1 cells, both of which require int erleukin-3 (IL-3) for their growth and survival. In the cells, KITG559 Or KITV814 were found to be constitutively phophorylated on tyrosine in the absence of stem cell factor (SCF) that is a ligand for KIT. Che mical cross-linking analysis showed that a substantial fraction of the phosphorylated KITG559 underwent dimerization even in the absence of SCF, whereas the phosphorylated KITV814 did not, suggesting the distin ct mechanisms underlying constitutive activation of KIT by G559 and V8 14 mutations. Furthermore, the cells expressing either KITG559 Or KITV 814 were found to show a factor-independent growth, whereas the cells expressing wildtype KIT (KITWT) proliferated in response to SCF as wel l as IL-3. Moreover, subcutaneous injection of Ba/F3 cells expressing KITG559 Or KITV814 into nude mice resulted in production of large tumo rs at all sites of the injection within 2 weeks, and all nude mice qui ckly succumbed to leukemia and died. These results suggest that, altho ugh the mechanisms underlying constitutive activation of KITG559 Or KI TV814 may be different, both of the activating mutations have a functi on to induce a factor-independent and tumorigenic phenotype. Also, the data of this study raise the possibility that the constitutively acti vating mutations of c-kit may play a causal role in development of hem atologic malignancies. (C) 1995 by The American Society of Hematology.