造血調節因子とそのレセプターによるシグナル伝達機構

1997 Fiscal Year Annual Research Report

• Project/Area Number: 06277101
• Research Institution: The University of Tokyo
• Principal Investigator: 新井 賢一 東京大学, 医科学研究所, 教授 (00012782)
• Co-Investigator(Kenkyū-buntansha): 宮島 篤 東京大学, 分子細胞生物学研究所, 教授 (50135232) ; 冨田 幹夫 埼玉県がんセンター, 主任研究員 (80142115) ; 松島 綱治 東京大学, 医学部, 教授 (50222427) ; 長田 重一 大阪大学, 医学部, 教授 (70114428) ; 高津 聖志 東京大学, 医科学研究所, 教授 (10107055)
• Keywords: 造血幹細胞 / サイトカイン / レセプター / インターロイキン / シグナル伝達 / プロテインキナーゼ / チロシンリン酸化 / 遺伝子破壊マウス

Research Abstract

新井らはGM-CSF受容体bc鎖の細胞内領域の全てのチロシン残基をフェニルアラニンに置換した変異体はGM-CSF依存性の細胞増殖シグナルの伝達、STAT5、MAPKの活性化を誘導することはできないがJak2を活性化することができ、しかもJak2特異的阻害剤AG-490によって阻害される抗アポトーシス効果をもつことを示した。宮島らは、IL-3/GM-CSFなどで活性化されるSTAT5の標的遺伝子を検索する過程でマウスOncostatin M(OSM)遺伝子をクローニングし、その受容体の解析から、ヒトとは異なり、マウスではOSMとLIFは受容体を共有しないことを明らかにした。OSMは生体の骨髄で強く発現しているが、骨髄由来の血球には作用しない。一方、OSMは胎生期のAGM領域での発現が認められ、AGMでの造血発生を強く促進することを明らかにした。高津らは、ヒトIL-5受容体a鎖にはJak2が、b鎖にはJak1が構成的に会合しており、IL-5刺激によって初めて両鎖の会合およびJak2の活性化が誘導され、さらにJak1,Shc,Vav,PI3kinaseの活性化が惹起されることを示した。長田らは、好中球に特異的に発現するZn-fingerをもつ転写因子mMZF-2のcDNAを単離し、その構造解析によりこの因子は814アミノ酸からなりN-末端部位には転写抑制部位、中央部には骨髄細胞特異的に作用する転写誘導部位、C-末端部位にはZn-fingerが13個からなるDNA結合部位があることを同定した。杉山らはWT1遺伝子を導入したマウス32Dc13細胞では、野生型では分化を誘導するG-CSFに反応して分化せずに増殖を始めること、さらにこの際Stat3a,Stat3bの活性化がみられたことによりWT1遺伝子の造血細胞におけるoncogenicな機能を検証した。富田らは、マウス骨髄性白血病細胞で発現させたGM-CSF受容体とLIF受容体のキメラ分子を解析し、LIF受容体細胞内領域のホモダイマーが細胞増殖停止と分化誘導のシグナルを伝達できること、及びSTAT3の活性化と分化誘導とに相関があることを示した。松島らは、ヒトCD34陽性、マウスLin陰性c-kit陽性造血幹細胞はCCR1を発現するが、IL8Rを発現しないこと、CCR1は赤芽球系前駆細胞にも発現し、MIP-1aがBFU-Eを抑制することを示し、in vitroでのマウス造血幹細胞からケモカイン受容体を発現する樹状突起細胞の分化誘導に成功し、さらにマウスLin陰性c-kit陽性造血幹細胞にそれぞれ独立に樹状突起細胞に分化するCD11b-/dullCD11c+とCD11+hiCD11c+の2つの樹状突起前駆細胞のサブセットがあることを示した。溝口らは彼らが樹立したヒト巨核球系細胞株Meg-Jにインドロカルバゾール系化合物であるK-252aを添加すると、著明に多倍体化するとともに成熟巨核球への分化傾向を示し、この多倍体化過程において、cdc2、サイクリンB1の発現およびcdc2活性は一過性の上昇が認められたのちに急速に減少したことを見い出し、M期を完遂しない巨核球の多倍体化過程においてcdc2キナーゼ活性は一過性に発現し、その特異的変動が多倍体化に関連していることを示した。

Research Products

• [Publications] Watanabe,S et.al: "Roles of JAK kinases in human GMCSF receptor signal transduction." J.Allergy Clinic Immunol.183-191 (1996)
• [Publications] Yasuda,Y.et.al: "A.Human granulocyto-macrophage colony-stimulating factor(hGM-CSF)induces inhibition of intrathymic T-cell development in hGM-CSF receptor transgenic mice." Blood. 89. 1349-1356 (1997)
• [Publications] Watanabe,S., Kubota,H: "Characterization of cis acting sequences and trans-acting signals regulating early growth response 1(egr-1)and c-fos promoters through the granulocyte-macrophage colony stimulating factor recepter in BA/F3 cells." Blood. 89. 1197-1206 (1997)
• [Publications] Liu,R., Itoh,T: "Activation of c-Jun N-Terminal Kinase(JNK)by Human Granulocyto Macrophage-Colony Stimulating Factor(hGM-CSF)in BA/F3 cells." Biochem.Biophys.Res.Commun. 234,3. 611-615 (1997)
• [Publications] Itoh,T., Liu,R.,: "Definition of the role of tyrosin residues of the common β subunit regulating multiple signaling pathways of GM CSF recepter." Mol.Cell.Biol. (in press).
• [Publications] Watanabe,S., Itoh,T: "Roles of JAK kinases in human GM-CSF receptor signals." International Association For Comparative Research on Leukemia and Related Diseases.
• [Publications] Yasue T: "Lynkinase is required for B cell triggering by CD38 ligation and IL-5 for IgGl production." Proc.Natl.Acad.Sci.USA. 94. 10307-10312 (1997)
• [Publications] Baba,M: "The role of CD40 ligation and cytokines in differentiation of mouse germinal center B cells towards antigen-specific antibody-forming cells:functional defects of germinal B cells of XID mice." Int.Immunol. 9. 1463-1473 (1997)
• [Publications] Ogata,N: "Signal transduction through human IL-5 receptor a chain in eosinophils." Int.Allergy Immunol. 114. 23-27 (1997)
• [Publications] Ogata,N: "An interleukin-5 receptor a subunit provides a binding site for IL-5 and functional signalling molecules as well." Blood. (in press). (1998)
• [Publications] Takahashi,K: "Effects of GM-CSF on the development and differentiation of CD5-positive macrophages and their derivation from a CD5-positive B cell lineage in mice." Am.J.Pathol. (in press). (1998)
• [Publications] Nagata,S: "Apoptosis by death factor." Cell. 88. 355-365 (1997)
• [Publications] Hueber,A: "Requirement for the CD95 reseptor-ligand pathway in c-myc-induced apoptosis." Science. 278. 1305-1309 (1997)
• [Publications] Kondo,T: "Essential roles of the Fas ligand in the development of hepatitis." Nature Med.3. 409-413 (1997)
• [Publications] Orita,T: "Binding of NF-Y Transcription Factor to One of Cis-Elements in the Myeloperoxidase Gene Promoter that Responds to G-CSF." J.Biol.Chem.272. 23216-23223 (1997)
• [Publications] Shimozaki,K: "Involvement of STAT3 in the granulocyte colony stimulating factor-induced differentiation of myeloid cells." J.Biol.Chem.272. 25184-25189 (1997)
• [Publications] Sakahira,H: "Clevage of CAD inhibitor in CAD activation and DNA degradation during apoptosis." Nature. 391. 96-99 (1998)
• [Publications] Enari,M.,: "A caspase-activated DNase that degrades DNA during apoptosis and its inhibitor ICAD." Nature. 391. 43-50 (1998)
• [Publications] Tanaka,M: "Down-regulation of Fas ligand by shedding." Nature Med. 4. 31-36 (1998)
• [Publications] Murai,K: "Myeloid-specific transcriptional activation by murine myeliod zinc finger protein-2." Proc.Matl.Acad.Sci.USA. (in press). (1998)
• [Publications] Inoue,K: "Aberrant overexpression of the Wilms tumor gene(WT1)in human leukemia." Blood. 89. 1405-1412 (1997)
• [Publications] Murata,Y: "The Wilms tumor suppressor gene WT1 unduces G1 arrest and apoptosis in myeloblastic leukemia M1 cells." FEBS Letters. 409. 41-45 (1997)
• [Publications] Kimura,T.: "Simultaneous activation of signals through gp130,c-kit,and interleukin-3 receptor promote a trilineage blood cell production in the absence of terminal acting lineage-specific factors." Blood. 90. 4767-4778 (1997)
• [Publications] Inoue,K: "Wilms'tumor gene(WT1)competes with differentiation-inducing signal in hematopoietic progenitor cells." Blood. (in press).
• [Publications] Sugiyama,H: "Wilms tumor gene(WT1)mRNA is equally expressed in blast cells from acute myeloid leukemia and normal CD34+progenitors(Response)." Blood. (in press).
• [Publications] Yamagami,T: "Suppression of Wilms'tumor gene(WT1)expression induces G2/M arrest in leukemic cells." Leukemia Research. (in press).
• [Publications] Zhang,Y: "Bifurcated dendritic cell differentiation in vitro from murine Lin-c-kit+bone marrow hematopoietic progenitor cells." Blood. in press (1998)
• [Publications] Wang,J-b.,: "Enhanced mobilization of hematopoietic progenitor cells by mouse macrophage inflammatory protein(MIP)-2 and granulocyte-colony stimulating factor in mice." J.Leukocyte Biol. 62. 503-509 (1997)
• [Publications] Zhang,Y: "Induction of dendritic cell deferentiation by GM-CSF,stem cell factor(SCF),and TNFa invitro from lineage phenotype(Lin-)c-kit+murine hematopoietic progenitor cells." Blood. 90. 4842-4853 (1997)
• [Publications] Su,S,-b.,: "Inhibition of immature erythroid progenitor cell proliferation by macrophage inflammatory protein-1a by interacting with C-C chemokine receptor,CCR-1." Blood. 90. 605-611 (1997)
• [Publications] Wada M,: "Molecular analysis of adenomatous polyposis coli gene in sarcomas,hematological malignancies and noncolonic,neoplastic tissues." J.Mol Med. 75. 139-144 (1997)
• [Publications] Teramura M: "Mechanism of action of antithymocyte globlin in the treatment of aplastic anemia:in vitro evidence for the presence of immunosuppressive mechanism." Br J Haematol. 96. 80-84 (1997)
• [Publications] Mori N: "Absence of microsatellite instability during the progression of chronic myelocytic leukemia." Leukemia. 151-152 (1997)
• [Publications] Masuda M: "Pure red cell aplasia with thymoma:evidence of T-cell clonal disorder." Am J Hematol. 54. 324-328 (1997)
• [Publications] Tsujimoto M: "Purification,CDNA Cloning,and Characterization of a New Serpin with Megakaryocyte Maturation Activity." J Bio Chem. 272. 15373-15380 (1997)
• [Publications] Mori N: "Allelotype analysis in the evolution of chronic myelocytic leukemia." Blood. 90・5. 2010-2014 (1997)
• [Publications] Yamada O: "Cyclophosphamide therapy for pure red cell aplasia associated with granular lymphocyte-proliferative disorders." Br J Haematol. 97. 392-399 (1997)
• [Publications] Ito K: "Development of a novel selective amplifier gene for controllableexpansion of transduced hematopoietic cells." Blood. 90・10. 3884-3892 (1997)
• [Publications] Takanashi M: "Proliferative effect of postapheresis platelet donor phasma on a megakaryoblatic leukemia cell line." Ann Hematol. 75. 149-153 (1997)
• [Publications] Teramura M: "Effect of thrombopoietin(c-Mpl ligand)alone and in combination with other hematopoietic growth factors on human megakaryocytopoiesis in serum-free cultures." Int J Hematology. 66. 373-381 (1997)
• [Publications] Tomida M: "Presence of mRNAs encoding the soluble D-factor/LIF receptor in human choriocarcinoma cells and production of the soluble receptor." Biochem Biophys Res Commun. 232. 427-431 (1997)
• [Publications] Tomida M: "Structure of the gene encoding the human differentiation-stimulating factor/leukemia inhibitory factor receptor." J Biochem. 120. 201-206 (1996)
• [Publications] Mukouyama,Y: "In vitro expansion of murine hematopoietic progenitors derived from the embryonic aorta gonad mesonephros region." Immunity. 8. 105-114 (1998)
• [Publications] Ichihara,M: "Oncostatin M and leukemia inhibitory factor do not utilize the same functional receptor in mice." Blood. 90. 165-173 (1997)
• [Publications] Arai,K: "Oxford Press" Mechanism of signal transduction(in press),
• [Publications] Takatsu,K: Interleukin-S.in"Growth factors and Cytokines in Health and Disease."vol.IIA(eds.Leroith),143-200 (1997)