Features of Ras activation by a mislocalized oncogenic tyrosine kinase: FLT3 ITD signals via K-Ras at the plasma membrane of Acute Myeloid Leukemia cells

Abstract

FLT3 ITD (FMS-like tyrosine kinase 3 with internal tandem duplication) is an important oncoprotein in Acute Myeloid Leukemia (AML). Owing to its constitutive kinase activity FLT3 ITD accumulates partially at endomembranes, a feature shared with other disease-associated, mutated receptor tyrosine kinases. Since Ras proteins also transit through endomembranes we have investigated the possible existence of an intracellular FLT3 ITD/Ras signaling pathway by comparing Ras signaling of FLT3 ITD with that of wild-type FLT3. Ligand stimulation activated both K- and N-Ras in cells expressing wild-type FLT3. Life-cell Ras-GTP imaging revealed ligand-induced Ras activation at the plasma membrane (PM). FLT3 ITD dependent constitutive activation of K-Ras and N-Ras was also observed primarily at the PM, supporting the view that the PM-resident pool of FLT3 ITD engaged the Ras/Erk pathway in AML cells. Accordingly, specific interference with FLT3 ITD/Ras signaling at the PM using PM-restricted dominant negative K-RasS17N potently inhibited cell proliferation and promoted apoptosis, corroborating that Ras signalling is crucial for FLT3 ITD dependent cell transformation and confirming that FLT3 ITD addresses PM-bound Ras despite its pronounced mislocalization to endomembranes.