title: Signaling of the ITK (interleukin 2-inducible T cell kinase)-SYK (spleen tyrosine kinase) fusion kinase is dependent on adapter SLP-76 and on the adapter function of the kinases SYK and ZAP70.
abstract: The inducible T cell kinase-spleen tyrosine kinase (ITK-SYK) oncogene consists of the Tec homology-pleckstrin homology domain of ITK and the kinase domain of SYK, and it is believed to be the cause of peripheral T cell lymphoma. We and others have recently demonstrated that this fusion protein is constitutively tyrosine-phosphorylated and is transforming both in vitro and in vivo. To gain a deeper insight into the molecular mechanism(s) underlying its activation and signaling, we mutated a total of eight tyrosines located in the SYK portion of the chimera into either phenylalanine or to the negatively charged glutamic acid. Although mutations in the interdomain-B region affected ITK-SYK kinase activity, they only modestly altered downstream signaling events. In contrast, mutations that were introduced in the kinase domain triggered severe impairment of downstream signaling. Moreover, we show here that SLP-76 is critical for ITK-SYK activation and is particularly required for the ITK-SYK-dependent phosphorylation of SYK activation loop tyrosines. In Jurkat cell lines, we demonstrate that expression of ITK-SYK fusion requires an intact SLP-76 function and significantly induces IL-2 secretion and CD69 expression. Furthermore, the SLP-76-mediated induction of IL-2 and CD69 could be further enhanced by SYK or ZAP-70, but it was independent of their kinase activity. Notably, ITK-SYK expression in SYF cells phosphorylates SLP-76 in the absence of SRC family kinases. Altogether, our data suggest that ITK-SYK exists in the active conformation state and is therefore capable of signaling without SRC family kinases or stimulation of the T cell receptor.
authors.affiliation: Department of Laboratory Medicine, Karolinska Institutet, Karolinska Hospital, Huddinge, SE 141 86 Stockholm, Sweden. alamdar.hussain@ki.se
authors.forename: Alamdar
authors.initials: A
authors.lastname: Hussain
authors.forename: Dara K
authors.initials: DK
authors.lastname: Mohammad
authors.forename: Manuela O
authors.initials: MO
authors.lastname: Gustafsson
authors.forename: Merve
authors.initials: M
authors.lastname: Uslu
authors.forename: Abdulrahman
authors.initials: A
authors.lastname: Hamasy
authors.forename: Beston F
authors.initials: BF
authors.lastname: Nore
authors.forename: Abdalla J
authors.initials: AJ
authors.lastname: Mohamed
authors.forename: C I Edvard
authors.initials: CI
authors.lastname: Smith
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D000945:Antigens, Differentiation, T-Lymphocyte / Q000378:metabolism; D015153:Blotting, Western; D019556:COS Cells; D005434:Flow Cytometry; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D019169:Jurkat Cells; D037181:Lectins, C-Type / Q000378:metabolism; D008213:Lymphocyte Activation; D009154:Mutation; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism; D010649:Phenylalanine / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D000072377:Syk Kinase; D013601:T-Lymphocytes / Q000378:metabolism; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism; D051746:ZAP-70 Protein-Tyrosine Kinase / Q000235:genetics / Q000378:metabolism*; D000099192:CD69 Antigens
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D018698:Glutamic Acid; D007376:Interleukin-2; D047908:Intracellular Signaling Peptides and Proteins; D037181:Lectins, C-Type; D015514:Oncogene Proteins, Fusion; D010649:Phenylalanine; D010750:Phosphoproteins; D011505:Protein-Tyrosine Kinases; D000072377:Syk Kinase; D014443:Tyrosine; D051746:ZAP-70 Protein-Tyrosine Kinase; D000099192:CD69 Antigens; C092947:SLP-76 signal Transducing adaptor proteins; C000606896:SYK protein, human; C497261:ZAP70 protein, human; C080813:emt protein-tyrosine kinase