TY - JOUR
T1 - Gain-of-function mutations in granulocyte colony–stimulating factor receptor (CSF3R) reveal distinct mechanisms of CSF3R activation
AU - Zhang, Haijiao
AU - Coblentz, Cody
AU - Watanabe-Smith, Kevin
AU - Means, Sophie
AU - Means, Jasmine
AU - Maxson, Julia E.
AU - Tyner, Jeffrey W.
N1 - Funding Information:
3Supported by the Leukemia and Lymphoma Society, the V Foundation for Cancer Research, Gabrielle’s Angel Foundation for Cancer Research, and NCI, National Institutes of Health Grants 5R00CA151457-04 and 1R01CA183947-01. To whom correspondence may be addressed: Oregon Health and Science University Knight Cancer Institute, 3181 S. W. Sam Jack-son Park Rd., BRB 553, Mailcode L592, Portland, OR 97239. Tel.: 503-494-9188; Fax: 503-494-3688; E-mail: tynerj@ohsu.edu.
Funding Information:
J. W. T. receives research support from Aptose, Array, AstraZeneca, Constellation, Genentech, Gilead, Incyte, Janssen, Seattle Genetics, Syros, and Takeda and is a member of the scientific advisory board for Leap Oncology. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. 2 Supported by an American Society of Hematology award and NCI, National Institutes of Health Grant R00CA190605-03. 3 Supported by the Leukemia and Lymphoma Society, the V Foundation for Cancer Research, Gabrielle’s Angel Foundation for Cancer Research, and NCI, National Institutes of Health Grants 5R00CA151457-04 and 1R01CA183947-01.
Publisher Copyright:
© 2018 Zhang et al. Published under exclusive license by The American Society for Biochemistry and Molecular Biology, Inc.
PY - 2018/5/11
Y1 - 2018/5/11
N2 - Granulocyte colony–stimulating factor (G-CSF or CSF3) and its receptor CSF3R regulate granulopoiesis, neutrophil function, and hematopoietic stem cell mobilization. Recent studies have uncovered an oncogenic role of mutations in the CSF3R gene in many hematologic malignancies. To find additional CSF3R mutations that give rise to cell transformation, we performed a cellular transformation assay in which murine interleukin 3 (IL-3)– dependent Ba/F3 cells were transduced with WT CSF3R plasmid and screened for spontaneous growth in the absence of IL-3. Any outgrowth clones were sequenced to identify CSF3R mutations with transformation capacity. We identified several novel mutations and determined that they transform cells via four distinct mechanisms: 1) cysteine- and disulfide bond–mediated dimerization (S581C); 2) polar, noncharged amino acid substitution at the transmembrane helix dimer interface at residue Thr-640; 3) increased internalization by a Glu-524 substitution that mimics a low G-CSF dose; and 4) hydrophobic amino acid substitutions in the membrane-proximal residues Thr-612, Thr-615, and Thr-618. Furthermore, the change in signaling activation was related to an altered CSF3R localization. We also found that CSF3R-induced STAT3 and ERK activations require CSF3R internalization, whereas STAT5 activation occurred at the cell surface. Cumulatively, we have expanded the regions of the CSF3R extracellular and transmembrane domains in which missense mutations exhibit leukemogenic capacity and have further elucidated the mechanistic underpinnings that underlie altered CSF3R expression, dimerization, and signaling activation.
AB - Granulocyte colony–stimulating factor (G-CSF or CSF3) and its receptor CSF3R regulate granulopoiesis, neutrophil function, and hematopoietic stem cell mobilization. Recent studies have uncovered an oncogenic role of mutations in the CSF3R gene in many hematologic malignancies. To find additional CSF3R mutations that give rise to cell transformation, we performed a cellular transformation assay in which murine interleukin 3 (IL-3)– dependent Ba/F3 cells were transduced with WT CSF3R plasmid and screened for spontaneous growth in the absence of IL-3. Any outgrowth clones were sequenced to identify CSF3R mutations with transformation capacity. We identified several novel mutations and determined that they transform cells via four distinct mechanisms: 1) cysteine- and disulfide bond–mediated dimerization (S581C); 2) polar, noncharged amino acid substitution at the transmembrane helix dimer interface at residue Thr-640; 3) increased internalization by a Glu-524 substitution that mimics a low G-CSF dose; and 4) hydrophobic amino acid substitutions in the membrane-proximal residues Thr-612, Thr-615, and Thr-618. Furthermore, the change in signaling activation was related to an altered CSF3R localization. We also found that CSF3R-induced STAT3 and ERK activations require CSF3R internalization, whereas STAT5 activation occurred at the cell surface. Cumulatively, we have expanded the regions of the CSF3R extracellular and transmembrane domains in which missense mutations exhibit leukemogenic capacity and have further elucidated the mechanistic underpinnings that underlie altered CSF3R expression, dimerization, and signaling activation.
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U2 - 10.1074/jbc.RA118.002417
DO - 10.1074/jbc.RA118.002417
M3 - Article
C2 - 29572350
AN - SCOPUS:85047061353
VL - 293
SP - 7387
EP - 7396
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 19
ER -