Inappropriate activation of oncogenic kinases at intracellular locations is frequently
observed in human cancers, but its effects on global signaling are incompletely understood.
Here, we show that the oncogenic mutant of Flt3 (Flt3-ITD), when localized at the
endoplasmic reticulum (ER), aberrantly activates STAT5 and upregulates its targets,
Pim-1/2, but fails to activate PI3K and MAPK signaling. Conversely, membrane targeting
of Flt3-ITD strongly activates the MAPK and PI3K pathways, with diminished phosphorylation
of STAT5. Global phosphoproteomics quantified 12,186 phosphorylation sites, confirmed
compartment-dependent activation of these pathways and discovered many additional
components of Flt3-ITD signaling. The differential activation of Akt and Pim kinases
by ER-retained Flt3-ITD helped to identify their putative targets. Surprisingly, we
find spatial regulation of tyrosine phosphorylation patterns of the receptor itself.
Thus, intracellular activation of RTKs by oncogenic mutations in the biosynthetic
route may exploit cellular architecture to initiate aberrant signaling cascades, thus
evading negative regulation.