title: CBL linker region and RING finger mutations lead to enhanced granulocyte-macrophage colony-stimulating factor (GM-CSF) signaling via elevated levels of JAK2 and LYN.
abstract: Juvenile myelomonocytic leukemia (JMML) is characterized by hypersensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF). SHP2, NF-1, KRAS, and NRAS are mutated in JMML patients, leading to aberrant regulation of RAS signaling. A subset of JMML patients harbor CBL mutations associated with 11q acquired uniparental disomy. Many of these mutations are in the linker region and the RING finger of CBL, leading to a loss of E3 ligase activity. We investigated the mechanism by which CBL-Y371H, a linker region mutant, and CBL-C384R, a RING finger mutant, lead to enhanced GM-CSF signaling. Expression of CBL mutants in the TF-1 cell line resulted in enhanced survival in the absence of GM-CSF. Cells expressing CBL mutations displayed increased phosphorylation of GM-CSF receptor βc subunit in response to stimulation, although expression of total GM-CSFR βc was lower. This suggested enhanced kinase activity downstream of GM-CSFR. JAK2 and LYN kinase expression is elevated in CBL-Y371H and CBL-C384R mutant cells, resulting in enhanced phosphorylation of CBL and S6 in response to GM-CSF stimulation. Incubation with the JAK2 inhibitor, TG101348, abolished the increased phosphorylation of GM-CSFR βc in cells expressing CBL mutants, whereas treatment with the SRC kinase inhibitor dasatinib resulted in equalization of GM-CSFR βc phosphorylation signal between wild type CBL and CBL mutant samples. Dasatinib treatment inhibited the elevated phosphorylation of CBL-Y371H and CBL-C384R mutants. Our study indicates that CBL linker and RING finger mutants lead to enhanced GM-CSF signaling due to elevated kinase expression, which can be blocked using small molecule inhibitors targeting specific downstream pathways.
authors.affiliation: Ontario Cancer Institute, Campbell Family Cancer Research Institute, Toronto, Ontario M5G 2M9, Canada.
authors.forename: Mojib
authors.initials: M
authors.lastname: Javadi
authors.forename: Terri D
authors.initials: TD
authors.lastname: Richmond
authors.forename: Kai
authors.initials: K
authors.lastname: Huang
authors.forename: Dwayne L
authors.initials: DL
authors.lastname: Barber
mesh_terms: D019943:Amino Acid Substitution; D002460:Cell Line; D053648:Cytokine Receptor Common beta Subunit / Q000235:genetics / Q000378:metabolism*; D000069439:Dasatinib; D015971:Gene Expression Regulation, Enzymologic; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects / Q000235:genetics; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D053614:Janus Kinase 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D054429:Leukemia, Myelomonocytic, Juvenile / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D020125:Mutation, Missense; D010766:Phosphorylation / Q000187:drug effects / Q000235:genetics; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D050721:Proto-Oncogene Proteins c-cbl / Q000096:biosynthesis* / Q000235:genetics; D011743:Pyrimidines / Q000494:pharmacology; D011759:Pyrrolidines / Q000494:pharmacology; D054829:RING Finger Domains / Q000235:genetics; D015398:Signal Transduction; D013449:Sulfonamides / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D019061:src-Family Kinases / Q000096:biosynthesis* / Q000235:genetics; D000096926:Benzenesulfonamides
chemical_list: D053648:Cytokine Receptor Common beta Subunit; D000069439:Dasatinib; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D053614:Janus Kinase 2; D047428:Protein Kinase Inhibitors; D050721:Proto-Oncogene Proteins c-cbl; D011743:Pyrimidines; D011759:Pyrrolidines; D013449:Sulfonamides; D013844:Thiazoles; D019061:src-Family Kinases; C528327:fedratinib; D000096926:Benzenesulfonamides; C507924:JAK2 protein, human; C060902:lyn protein-tyrosine kinase; C067419:CBL protein, human
keywords: CBL; Cytokine; E3 Ubiquitin Ligase; GM-CSF; JAK Kinase; Juvenile Myelomonocytic Leukemia; SRC; Signal Transduction