Protein kinase B (c-akt) regulates hematopoietic lineage choice decisions during myelopoiesis.

Hematopoiesis is a highly regulated process resulting in the formation of all blood lineages. Aberrant regulation of phosphatidylinositol-3-kinase (PI3K) signaling has been observed in hematopoietic malignancies, suggesting that regulated PI3K signaling is critical for regulation of blood cell production. An ex vivo differentiation system was used to investigate the role of PI3K and its downstream effector, protein kinase B (PKB/c-akt) in myelopoiesis. PI3K activity was essential for hematopoietic progenitor survival. High PKB activity was found to promote neutrophil and monocyte development, while, conversely, reduction of PKB activity was required to induce optimal eosinophil differentiation. In addition, transplantation of beta2-microglobulin (-/-) NOD/SCID mice with CD34(+) cells ectopically expressing constitutively active PKB resulted in enhanced neutrophil and monocyte development, whereas ectopic expression of dominant-negative PKB induced eosinophil development in vivo. Inhibitory phosphorylation of C/EBPalpha on Thr222/226 was abrogated upon PKB activation in hematopoietic progenitors. Ectopic expression of a nonphosphorylatable C/EBPalpha mutant inhibited eosinophil differentiation ex vivo, whereas neutrophil development was induced, demonstrating the importance of PKB-mediated C/EBPalpha phosphorylation in regulation of granulopoiesis. These results identify an important novel role for PKB in regulation of cell fate choices during hematopoietic lineage commitment.

[1]  J. Blenis,et al.  An inhibitor of mTOR reduces neoplasia and normalizes p70/S6 kinase activity in Pten+/− mice , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[2]  O. MacDougald,et al.  Glycogen Synthase Kinase 3 Is an Insulin-Regulated C/EBPα Kinase , 1999, Molecular and Cellular Biology.

[3]  Hong Liu,et al.  Involvement of Protein Kinase Cε in the Negative Regulation of Akt Activation Stimulated by Granulocyte Colony-Stimulating Factor1 , 2006, The Journal of Immunology.

[4]  D. Tenen,et al.  Phosphorylation of C/EBPα Inhibits Granulopoiesis , 2004, Molecular and Cellular Biology.

[5]  C. Potter,et al.  Drosophila Tsc1 Functions with Tsc2 to Antagonize Insulin Signaling in Regulating Cell Growth, Cell Proliferation, and Organ Size , 2001, Cell.

[6]  P. Piccaluga,et al.  Frequent elevation of Akt kinase phosphorylation in blood marrow and peripheral blood mononuclear cells from high-risk myelodysplastic syndrome patients , 2006, Leukemia.

[7]  T. Graf,et al.  Lineage commitment of transformed haematopoietic progenitors is determined by the level of PKC activity. , 1996, The EMBO journal.

[8]  D. Guertin,et al.  Rictor, a Novel Binding Partner of mTOR, Defines a Rapamycin-Insensitive and Raptor-Independent Pathway that Regulates the Cytoskeleton , 2004, Current Biology.

[9]  C. Sugimoto,et al.  PI3K/Akt and apoptosis: size matters , 2003, Oncogene.

[10]  B. Burgering,et al.  Protein kinase B (c-Akt) in phosphatidylinositol-3-OH kinase signal transduction , 1995, Nature.

[11]  J. Lammers,et al.  Signal transducer and activator of transcription 5a (STAT5a) is required for eosinophil differentiation of human cord blood-derived CD34+ cells. , 2003, Blood.

[12]  W. V. van Blitterswijk,et al.  Protein Kinase Cζ Is a Negative Regulator of Protein Kinase B Activity* , 1999, The Journal of Biological Chemistry.

[13]  D. Tenen,et al.  Phosphorylation of C/EBPalpha inhibits granulopoiesis. , 2004, Molecular and cellular biology.

[14]  I. Weissman,et al.  Identification of Clonogenic Common Lymphoid Progenitors in Mouse Bone Marrow , 1997, Cell.

[15]  Hong Yang,et al.  ANG II stimulation of neuritogenesis involves protein kinase B in brain neurons. , 2002, American journal of physiology. Regulatory, integrative and comparative physiology.

[16]  M. Birnbaum,et al.  Akt1/PKBα Is Required for Normal Growth but Dispensable for Maintenance of Glucose Homeostasis in Mice* , 2001, The Journal of Biological Chemistry.

[17]  D. Tenen,et al.  CCAAT/Enhancer Binding Protein α Is a Regulatory Switch Sufficient for Induction of Granulocytic Development from Bipotential Myeloid Progenitors , 1998, Molecular and Cellular Biology.

[18]  I. Weissman,et al.  The long-term repopulating subset of hematopoietic stem cells is deterministic and isolatable by phenotype. , 1994, Immunity.

[19]  D. Tenen,et al.  Absence of granulocyte colony-stimulating factor signaling and neutrophil development in CCAAT enhancer binding protein alpha-deficient mice. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[20]  N. Hay,et al.  The Akt-mTOR tango and its relevance to cancer. , 2005, Cancer cell.

[21]  R. Loewith,et al.  Mammalian TOR complex 2 controls the actin cytoskeleton and is rapamycin insensitive , 2004, Nature Cell Biology.

[22]  Xi C. He,et al.  PTEN maintains haematopoietic stem cells and acts in lineage choice and leukaemia prevention , 2006, Nature.

[23]  E. Solary,et al.  Essential role for the p110delta isoform in phosphoinositide 3-kinase activation and cell proliferation in acute myeloid leukemia. , 2005, Blood.

[24]  I. Weissman,et al.  A clonogenic common myeloid progenitor that gives rise to all myeloid lineages , 2000, Nature.

[25]  K. Kaibuchi,et al.  Rho GTPase/Rho Kinase Negatively Regulates Endothelial Nitric Oxide Synthase Phosphorylation through the Inhibition of Protein Kinase B/Akt in Human Endothelial Cells , 2002, Molecular and Cellular Biology.

[26]  E. Solary,et al.  Essential role for the p110δ isoform in phosphoinositide 3-kinase activation and cell proliferation in acute myeloid leukemia , 2005 .

[27]  G. Thomas,et al.  Disruption of the Mouse mTOR Gene Leads to Early Postimplantation Lethality and Prohibits Embryonic Stem Cell Development , 2004, Molecular and Cellular Biology.

[28]  M. Raffeld,et al.  Human mesenchymal stem cells exert potent antitumorigenic effects in a model of Kaposi's sarcoma , 2006, The Journal of experimental medicine.

[29]  W L Stanford,et al.  Function of PI3Kgamma in thymocyte development, T cell activation, and neutrophil migration. , 2000, Science.

[30]  R. Hresko,et al.  mTOR·RICTOR Is the Ser473 Kinase for Akt/Protein Kinase B in 3T3-L1 Adipocytes* , 2005, Journal of Biological Chemistry.

[31]  K. Okkenhaug,et al.  Impaired B and T Cell Antigen Receptor Signaling in p110δ PI 3-Kinase Mutant Mice , 2002, Science.

[32]  Andrius Kazlauskas,et al.  The protein kinase encoded by the Akt proto-oncogene is a target of the PDGF-activated phosphatidylinositol 3-kinase , 1995, Cell.

[33]  Hong Wu,et al.  Enhanced sensitivity of PTEN-deficient tumors to inhibition of FRAP/mTOR , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[34]  J. Stock,et al.  Severe diabetes, age-dependent loss of adipose tissue, and mild growth deficiency in mice lacking Akt2/PKB beta. , 2003, The Journal of clinical investigation.

[35]  N. Sonenberg,et al.  Upstream and downstream of mTOR. , 2004, Genes & development.

[36]  J. Lammers,et al.  Signal transducer and activator of transcription 5 a ( STAT 5 a ) is required for eosinophil differentiation of human cord blood – derived CD 34 cells , 2002 .

[37]  J. Woodgett,et al.  Negative regulation of phosphatidylinositol 3-kinase and Akt signalling pathway by PKC. , 2003, Cellular signalling.

[38]  Jennifer Skeen,et al.  Dwarfism, impaired skin development, skeletal muscle atrophy, delayed bone development, and impeded adipogenesis in mice lacking Akt1 and Akt2. , 2003, Genes & development.

[39]  J. Myklebust,et al.  Activation of phosphatidylinositol 3-kinase is important for erythropoietin-induced erythropoiesis from CD34(+) hematopoietic progenitor cells. , 2002, Experimental hematology.

[40]  R. Moon,et al.  Glycogen synthase kinase-3 is an in vivo regulator of hematopoietic stem cell repopulation , 2006, Nature Medicine.

[41]  M. Hall,et al.  TOR Signaling in Growth and Metabolism , 2006, Cell.

[42]  O. MacDougald,et al.  Glycogen synthase kinase 3 is an insulin-regulated C/EBPalpha kinase. , 1999, Molecular and cellular biology.

[43]  E. Wilder,et al.  Cell-autonomous regulation of cell and organ growth in Drosophila by Akt/PKB , 1999, Nature Cell Biology.

[44]  E. Furth,et al.  Regulation of pancreatic beta-cell growth and survival by the serine/threonine protein kinase Akt1/PKBalpha. , 2001, Nature medicine.

[45]  J. Lammers,et al.  Differential regulation of granulopoiesis by the basic helix-loop-helix transcriptional inhibitors Id1 and Id2. , 2005, Blood.

[46]  I. Roninson,et al.  Growth retardation and increased apoptosis in mice with homozygous disruption of the Akt1 gene. , 2001, Genes & development.

[47]  F. Alt,et al.  Impaired B cell development and proliferation in absence of phosphoinositide 3-kinase p85alpha. , 1999, Science.