Translocation of Akt/PKB to the nucleus of osteoblast‐like MC3T3‐E1 cells exposed to proliferative growth factors

FEBS Letters

Casto, Riccardo²  Martelli, Alberto M¹  Borgatti, Paola⁴  Capitani, Silvano⁴  Bellacosa, Alfonso³  Massari, Leo²  Neri, Luca M⁴ 

[1]Dipartimento di Morfologia Umana Normale, Università di Trieste, via Manzoni 16, 34138 Trieste, Italy

[2]Dipartimento di Scienze Biomediche e Terapie Avanzate, Sezione di Clinica Ortopedica, Università di Ferrara, Corso della Giovecca 203, 44100 Ferrara, Italy

[3]Division of Population Science, Fox Chase Cancer Center, 7701 Burholme Avenue, Philadelphia, PA 19111, USA

[4]Dipartimento di Morfologia ed Embriologia, Sezione di Anatomia Umana, Università di Ferrara, via Fossato di Mortara 66, 44100 Ferrara, Italy

关键词: Akt/PKB;    Nucleus;    Signal transduction;    Platelet-derived growth factor;    Insulin-like growth factor;    MC3T3-E1 osteoblast-like cell;    PKB;    protein kinase B;    PKC;    protein kinase C;    PKA;    protein kinase A;    PI3K;    phosphatidylinositol 3-kinase;    PDK1;    phospholipid-dependent kinase 1;    PtdIns(3;    4;    5)P3;    phosphatidylinositol-3;    4;    5-trisphosphate;    DAG;    diacylglycerol;    IGF-I;    insulin-like growth factor;    PDGF;    platelet-derived growth factor;    EGF;    epidermal growth factor;    FGF;    fibroblast growth factor;    NGS;    normal goat serum;    FCS;    fetal calf serum;    BSA;    bovine serum albumin;    D-MEM;    Dulbecco's modified minimum essential medium;    PBS;    phosphate-buffered saline;    CLSM;    confocal laser scanning microscope;    PMSF;    phenylmethylsulfonyl fluoride;    EGTA;    ethylene glycol-bis(β-aminoethyl ether) N;    N;    N′;    N′-tetraacetic acid;    DTT;    dithiothreitol;    5-BrdU;    5-bromodeoxyuridine;   

DOI  :  10.1016/S0014-5793(00)01758-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

An active phosphatidylinositol 3-kinase (PI3K) has been shown in nuclei of different cell types. The products of this enzyme, i.e. inositides phosphorylated in the D3 position of the inositol ring, may act as second messengers themselves. Nuclear PI3K translocation has been demonstrated to be related to an analogous translocation of a PtdIns(3,4,5)P₃ activated PKC, the ζ isozyme. We have examined the issue of whether or not in the osteoblast-like clonal cell line MC3T3-E1 there may be observed an insulin-like growth factor-I- (IGF-I) and platelet-derived growth factor- (PDGF) dependent nuclear translocation of an active Akt/PKB. Western blot analysis showed a maximal nuclear translocation after 20 min of IGF-I stimulation or after 30 min of PDGF treatment. Both growth factors increased rapidly and transiently the enzyme activity of immunoprecipitable nuclear Akt/PKB on a similar time scale and after 60 min the values were slightly higher than the basal levels. Enzyme translocation was blocked by the specific PI3K inhibitor, LY294002, as well as cell entry into S-phase. Confocal microscopy showed an evident increase in immunostaining intensity in the nuclear interior after growth factor treatment but no changes in the subcellular distribution of Akt/PKB when a LY294002 pre-treatment was administered to the cells. These findings strongly suggest that the intranuclear translocation of Akt/PKB is an important step in signalling pathways that mediate cell proliferation.

【 授权许可】

Unknown