Constitutively activated FGFR3 mutants signal through PLCγ-dependent and -independent pathways for hematopoietic transformation

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Abstract
Ectopic expression of fibroblast growth factor receptor 3 (FGFR3) associated with t(4;14) has been implicated in the pathogenesis of human multiple myeloma. Some t(4;14) patients have activating mutations of FGFR3, of which a minority are K650E (thanatophoric dysplasia type II [TDII]). To investigate the role of autophosphorylated tyrosine residues in FGFR3 signal transduction and transformation, we characterized a series of FGFR3 TDII mutants with single or multiple Y→F substitutions. Phenylalanine substitution of Y760, essential for phospholipase Cγ (PLCγ) binding and activation, significantly attenuated FGFR3 TDII–mediated PLCγ activation, as well as transformation in Ba/F3 cells and a murine bone marrow transplant leukemia model. In contrast, single substitution of Y577, Y724, or Y770 had minimal to moderate effects on TDII-dependent transformation. Substitution of all 4 non–activation loop tyrosine residues significantly attenuated, but did not abolish, TDII transforming activity. Similar observations were obtained in the context of a constitutively activated fusion TEL-FGFR3 associated with t(4;12)(p16;p13) peripheral T-cell lymphomas. Moreover, 2 independent EμSR-FGFR3 TDII transgenic mouse lines developed a pro-B-cell lymphoma, and PLCγ was highly activated in primary lymphoma cells as assessed by tyrosine phosphorylation. These data indicate that engagement of multiple signaling pathways, including PLCγ-dependent and PLCγ-independent pathways, is required for full hematopoietic transformation by constitutively activated FGFR3 mutants.