VASCULAR ENDOTHELIAL-CELL GROWTH-FACTOR PROMOTES TYROSINE PHOSPHORYLATION OF MEDIATORS OF SIGNAL-TRANSDUCTION THAT CONTAIN SH2 DOMAINS - ASSOCIATION WITH ENDOTHELIAL-CELL PROLIFERATION
VASCULAR ENDOTHELIAL-CELL GROWTH-FACTOR PROMOTES TYROSINE PHOSPHORYLATION OF MEDIATORS OF SIGNAL-TRANSDUCTION THAT CONTAIN SH2 DOMAINS - ASSOCIATION WITH ENDOTHELIAL-CELL PROLIFERATION
复制标题
DOI：
10.1074/jbc.270.12.6729
复制
发表时间：
1995-03-24
影响因子：
4.8
通讯作者：
DONNER, DB
中科院分区：
文献类型：
--
作者：
GUO, DQ;JIA, Q;DONNER, DB
血管内皮细胞生长因子（VEGF）是一种内皮细胞特异性的有丝分裂原，在血管生成中起重要作用，可促进牛主动脉内皮细胞（BAEC）中至少11种蛋白质的酪氨酸磷酸化。蛋白质免疫沉淀的对照和VEGF刺激BAEC与抗血清磷脂酶C-γ（PLC-γ）的裂解物通过SDS-聚丙烯酰胺凝胶电泳进行分级，并转移到Immobilon-P。蛋白质印迹与抗血清磷酸酪氨酸的评价表明，PLC-γ和两种蛋白质（100和85 kDa），与PLC-γ磷酸化响应VEGF。通过使用对含有SH 2结构域的信号转导的其他介质具有特异性的抗血清进行免疫沉淀，证明了VEGF促进磷脂酰肌醇3-激酶、Ras GT3活化蛋白（GAP）和致癌衔接蛋白NcK的磷酸化。与VEGF受体Flt-1和Flk-1/KDR一致的M（r）蛋白在刺激的细胞中也被酪氨酸磷酸化，酪氨酸磷酸化的Nck、PLC-γ和两种GAP相关蛋白p190和p62在VEGF刺激的BAEC的GAP免疫沉淀物中，酪氨酸磷酸化的NcK在磷脂酰肌醇3-激酶免疫沉淀物中，这些观察结果表明，VEGF促进VEGF受体与含有SH 2结构域的蛋白质的多聚体聚集体的形成，并激活各种信号传导途径。酪氨酸激酶抑制剂genistein可抑制VEGF促进的内皮细胞增殖和含SH 2结构域信号分子的酪氨酸磷酸化。
Vascular endothelial cell growth factor (VEGF), an endothelial cell-specific mitogen that plays an important role in angiogenesis, promotes the tyrosine phosphorylation of at least 11 proteins in bovine aortic endothelial cells (BAEC). Proteins immunoprecipitated fr om lysates of control- and VEGF stimulated BAEC with antisera to phospholipase C-gamma (PLC-gamma) were fractionated by SDS-polyacrylamide gel electrophoresis and transferred to Immobilon-P. Evaluation of the Western blots with antisera to phosphotyrosine demonstrated that PLC-gamma and two proteins (100 and 85 kDa) that associate with PLC-gamma were phosphorylated in response to VEGF. By using antisera specific to other mediators of signal transduction that contain SH2 domains for immunoprecipitation, it was demonstrated that VEGF promotes phosphorylation of phosphatidylinositol 3-kinase, Ras GTPase activating protein (GAP), and the oncogenic adaptor protein NcK. Proteins of M(r) consistent with the VEGF receptors Flt-1 and Flk-1/KDR were also tyrosine phosphorylated in stimulated cells, Tyrosine-phosphorylated Nck, PLC-gamma, and two GAP-associated proteins, p190 and p62, were in GAP immunoprecipitates of VEGF-stimulated BAEC, and tyrosine-phosphorylated NcK was in phosphatidylinositol 3-kinase immunoprecipitates, These observations suggest that VEGF promotes formation of multimeric aggregates of VEGF receptors with proteins that contain SH2 domains and activate various signaling pathways. VEGF-promoted proliferation of endothelial cells and tyrosine phosphorylation of SH2 domain containing signaling molecules were inhibited by the tyrosine kinase inhibitor genistein.