Loss of SHIP and CIS recruitment to the granulocyte colony-stimulating factor receptor contribute to hyperproliferative responses in severe congenital neutropenia/acute myelogenous leukemia.
Mutations in the G-CSF receptor (G-CSFR) in patients with severe congenital neutropenia (SCN) are postulated to contribute to transformation to acute myelogenous leukemia (AML). These mutations result in defective receptor internalization and sustained cellular activation, suggesting a loss of negative signaling by the G-CSFR. In this paper we investigated the roles of SHIP and cytokine-inducible Src homology 2 protein (CIS) in down-modulating G-CSFR signals and demonstrate that loss of their recruitment as a consequence of receptor mutations leads to aberrant signaling. We show that SHIP binds to phosphopeptides corresponding to Tyr744 and Tyr764 in the G-CSFR and that Tyr764 is required for in vivo phosphorylation of SHIP and the formation of SHIP/Shc complexes. Cells expressing a G-CSFR form lacking Tyr764 exhibited hypersensitivity to G-CSF and enhanced proliferation, but to a lesser degree than observed with the most common mutant G-CSFR form in patients with SCN/AML, prompting us to investigate whether suppressor of cytokine signaling proteins also down-modulate G-CSFR signals. G-CSF was found to induce the expression of CIS and of CIS bound to phosphopeptides corresponding to Tyr729 and Tyr744 of the G-CSFR. The expression of CIS was prolonged in cells with the SCN/AML mutant G-CSFR lacking Tyr729 and Tyr744, which also correlated with increased G-CSFR expression. These findings suggest that SHIP and CIS interact with distal phosphotyrosine residues in the G-CSFR to negatively regulate G-CSFR signaling by limiting proliferation and modulating surface expression of the G-CSFR, respectively. Novel therapeutic approaches targeting inhibitory pathways that limit G-CSFR signaling may have promise in the treatment of patients with SCN/AML.
Bone Marrow Transplantation Program, The Arthur G. James Cancer Hospital, Ohio State University, Columbus 43210, USA.
Melissa G
MG
Hunter
Anand
A
Jacob
Lynn C
LC
O'donnell
Amanda
A
Agler
Lawrence J
LJ
Druhan
K Mark
KM
Coggeshall
Belinda R
BR
Avalos
D002352:Carrier Proteins / Q000032:analysis; D002455:Cell Division; D002470:Cell Survival; D004268:DNA-Binding Proteins / Q000378:metabolism; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D017874:Immediate-Early Proteins / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins; D015470:Leukemia, Myeloid, Acute / Q000473:pathology*; D008894:Milk Proteins / Q000378:metabolism; D009503:Neutropenia / Q000151:congenital* / Q000473:pathology*; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases / Q000502:physiology*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000502:physiology*; D012097:Repressor Proteins / Q000032:analysis; D050799:STAT5 Transcription Factor; D015398:Signal Transduction; D000071222:Suppressor of Cytokine Signaling 1 Protein; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000032:analysis
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D016179:Granulocyte Colony-Stimulating Factor; D017874:Immediate-Early Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008894:Milk Proteins; D000072183:Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; D010744:Phosphoric Monoester Hydrolases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D012097:Repressor Proteins; D050799:STAT5 Transcription Factor; D000071222:Suppressor of Cytokine Signaling 1 Protein; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C108944:SOCS1 protein, human; C106644:SOCS3 protein, human; C098706:INPPL1 protein, human