title: The lymphoma-associated fusion tyrosine kinase ITK-SYK requires pleckstrin homology domain-mediated membrane localization for activation and cellular transformation.
abstract: ITK-SYK, a novel fusion tyrosine kinase (FTK) resulting from a recurrent t(5;9)(q33;q22), was recently identified in a poorly understood subset of peripheral T-cell lymphomas. However, the biochemical and functional properties of ITK-SYK are unknown. Here we demonstrate that ITK-SYK is a catalytically active tyrosine kinase that is sensitive to an established inhibitor of SYK. The expression of ITK-SYK, but not SYK, transformed NIH3T3 cells, inducing loss of contact inhibition and formation of anchorage-independent colonies in soft agar, in a kinase activity-dependent manner. ITK-SYK is unusual among FTKs in having an N-terminal phosphatidylinositol 3,4,5-trisphosphate-binding pleckstrin homology (PH) domain. Introduction of a well characterized loss-of-function mutation (R29C) into the PH domain of ITK-SYK inhibited its phosphorylation, markedly reduced its catalytic activity, and abrogated its ability to activate the ERK signaling pathway and to transform NIH3T3 cells. Although ITK-SYK was membrane-associated, ITK-SYK-R29C was not. However, each of these properties could be recovered by retargeting ITK-SYK-R29C back to the plasma membrane by the addition of an N-terminal myristylation sequence. Consistent with a model in which ITK-SYK requires PH domain-mediated binding to phosphatidylinositol 3,4,5-trisphosphate generated by phosphatidylinositol 3-kinase (PI3K), ITK-SYK activity was reduced by pharmacological inhibition of PI3K and increased by co-expression with a constitutively active form of PI3K. Together, these findings identify ITK-SYK as an active, transforming FTK dependent upon PH domain-mediated membrane localization, identify a novel mechanism for activation of an oncogenic FTK, and suggest ITK-SYK as a rational therapeutic target for t(5;9)(q33;q22)-positive lymphomas.
authors.affiliation: Division of Molecular Histopathology, Department of Pathology, University of Cambridge, Cambridge CB2 2QQ, United Kingdom.
authors.forename: Sue
authors.initials: S
authors.lastname: Rigby
authors.forename: Yuanxue
authors.initials: Y
authors.lastname: Huang
authors.forename: Berthold
authors.initials: B
authors.lastname: Streubel
authors.forename: Andreas
authors.initials: A
authors.lastname: Chott
authors.forename: Ming-Qing
authors.initials: MQ
authors.lastname: Du
authors.forename: Suzanne D
authors.initials: SD
authors.lastname: Turner
authors.forename: Chris M
authors.initials: CM
authors.lastname: Bacon
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002471:Cell Transformation, Neoplastic; D006801:Humans; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D019169:Jurkat Cells; D016411:Lymphoma, T-Cell, Peripheral / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051379:Mice; D018613:Microscopy, Confocal; D020125:Mutation, Missense; D041681:NIH 3T3 Cells; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D000072377:Syk Kinase; D014162:Transfection
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D015514:Oncogene Proteins, Fusion; D018129:Phosphatidylinositol Phosphates; C060974:phosphatidylinositol 3,4,5-triphosphate; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C000606897:Syk protein, mouse; C080813:emt protein-tyrosine kinase