title: Phosphoproteomic analysis of anaplastic lymphoma kinase (ALK) downstream signaling pathways identifies signal transducer and activator of transcription 3 as a functional target of activated ALK in neuroblastoma cells.
abstract: Activation of the anaplastic lymphoma kinase (ALK) receptor tyrosine kinase is a key oncogenic mechanism in a growing number of tumor types. In the majority of cases, ALK is activated by fusion with a dimerizing partner protein as a result of chromosomal translocation events, most studied in the case of the nucleophosmin-ALK and echinoderm microtubule-associated protein-like 4-ALK oncoproteins. It is now also appreciated that the full-length ALK receptor can be activated by point mutations and by deletions within the extracellular domain, such as those observed in neuroblastoma. Several studies have employed phosphoproteomics approaches to find substrates of ALK fusion proteins. In this study, we used MS-based phosphotyrosine profiling to characterize phosphotyrosine signaling events associated with the full-length ALK receptor. A number of previously identified and novel targets were identified. One of these, signal transducer and activator of transcription 3 (STAT3), has previously been observed to be activated in response to oncogenic ALK signaling, but the significance of this in signaling from the full-length ALK receptor has not been explored further. We show here that activated ALK robustly activates STAT3 on Tyr705 in a number of independent neuroblastoma cell lines. Furthermore, knockdown of STAT3 by RNA interference resulted in a reduction in myelocytomatosis neuroblastom (MYCN) protein levels downstream of ALK signaling. These observations, together with a decreased level of MYCN and inhibition of neuroblastoma cell growth in the presence of STAT3 inhibitors, suggest that activation of STAT3 is important for ALK signaling activity in neuroblastoma.
authors.affiliation: Department of Molecular Biology, Umeå University, Sweden.
authors.forename: Kamaraj
authors.initials: K
authors.lastname: Sattu
authors.forename: Falko
authors.initials: F
authors.lastname: Hochgräfe
authors.forename: Jianmin
authors.initials: J
authors.lastname: Wu
authors.forename: Ganesh
authors.initials: G
authors.lastname: Umapathy
authors.forename: Christina
authors.initials: C
authors.lastname: Schönherr
authors.forename: Kristina
authors.initials: K
authors.lastname: Ruuth
authors.forename: Damini
authors.initials: D
authors.lastname: Chand
authors.forename: Barbara
authors.initials: B
authors.lastname: Witek
authors.forename: James
authors.initials: J
authors.lastname: Fuchs
authors.forename: Pui-Kai
authors.initials: PK
authors.lastname: Li
authors.forename: Fredrik
authors.initials: F
authors.lastname: Hugosson
authors.forename: Roger J
authors.initials: RJ
authors.lastname: Daly
authors.forename: Ruth H
authors.initials: RH
authors.lastname: Palmer
authors.forename: Bengt
authors.initials: B
authors.lastname: Hallberg
mesh_terms: D000077548:Anaplastic Lymphoma Kinase; D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D049109:Cell Proliferation; D006801:Humans; D047468:Immunoprecipitation; D008156:Luciferases; D009447:Neuroblastoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D016716:PC12 Cells; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D020543:Proteome / Q000032:analysis*; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050796:STAT3 Transcription Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D010750:Phosphoproteins; D020543:Proteome; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D050796:STAT3 Transcription Factor; D019000:Phosphotyrosine; D008156:Luciferases; C000626173:ALK protein, human; C000626175:Alk protein, rat; D000077548:Anaplastic Lymphoma Kinase; D020794:Receptor Protein-Tyrosine Kinases
keywords: SHP-2; anaplastic lymphoma kinase; cancer; neuroblastoma; signal transducer and activator of transcription 3 (STAT3)
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