HEMATOPOIESIS Phosphatidylinositol 3 kinase contributes to the transformation of hematopoietic cells by the D816V c-Kit mutant

Stem cell factor (SCF) binds the receptor tyrosine kinase c-Kit and is critical for normal hematopoiesis. Substitution of va-line for aspartic acid 816 (D816V) constitu-tively actives human c-Kit, and this muta-tion is found in patients with mastocytosis, leukemia, and germ cell tumors. Immortal-ized murine progenitor cells (MIHCs) trans-duced with wild-type c-Kit proliferate in re-sponse to SCF, whereas cells expressing D816V c-Kit (MIHC-D816V) are factor-inde-pendent and tumorigenic. However, the mechanisms mediating transformation by D816V c-Kit are unknown. The objective of this study was to identify signaling compo-nents that contribute to D816V c-Kit–medi-ated transformation. SCF stimulates asso-ciation of p85PI3K with phosphorylated tyrosine 721 of wild-type c-Kit. Phosphatidyl-inositol 3 kinase (PI3K) subsequently con-tributes to the activation of Akt and Jnks. In contrast, these studies demonstrated that the D816V c-Kit mutant was constitutively associatedwithphosphorylatedp85PI3K,and, downstream of PI3K, Jnk 1 and Jnk 2 were activated butAkt was not. Interestingly, Erks 1 and 2 were not constitutively activated by D816V c-Kit. Thus, D816V c-Kit maintains the activity of PI3K but not of all signaling pathways activated by wild-type c-Kit. Further, all pathways downstream of PI3K are not constitutively active in MIHC-D816V cells. Studies with a PI3K inhibitor and D816V/Y721F c-Kit, a mutant incapable of recruiting PI3K, indicate that constitutive activation of PI3K through direct recruit-ment by D816V c-Kit plays a role in factor-independent growth of MIHC and is critical for tumorigenicity. (Blood. 2001;98