Fukuda Seiji, Foster Richard G, Porter Scott B, Pelus Louis M
Department of Microbiology and Immunology and the Walther Oncology Center, Indiana University School of Medicine; and the Walther Cancer Institute, Indianapolis, IN 46202, USA.
Blood. 2002 Oct 1;100(7):2463-71. doi: 10.1182/blood.V100.7.2463.
The inhibitor of the apoptosis protein (IAP) survivin is expressed in proliferating cells such as fetal tissues and cancers. We previously reported that survivin is expressed and growth factor regulated in normal adult CD34(+) cells. Herein, we examined survivin expression in CD34(+) cells before and after cell cycle entry and demonstrate a role for survivin in cell cycle regulation and proliferation. Analysis of known human IAPs revealed that only survivin is cytokine regulated in CD34(+) cells. Survivin expression is coincident with cell cycle progression. Up-regulation of survivin by thrombopoietin (Tpo), Flt3 ligand (FL), and stem cell factor (SCF) occurred in underphosphorylated-retinoblastoma protein (Rb)(positive), Ki-67(negative), and cyclin D(negative) CD34(+) cells. Quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) and multivariate flow cytometry demonstrated that Tpo, SCF, and FL increase survivin mRNA and protein in quiescent G(0) CD34(+) cells without increasing Ki-67 expression, indicating that cytokine-stimulated up-regulation of survivin in CD34(+) cells occurs during G(0), before cells enter G(1). Selective inhibition of the PI3-kinase/AKT and mitogen-activated protein kinase (MAPK(p42/44)) pathways blocked survivin up-regulation by growth factors before arresting cell cycle. Retrovirus transduction of survivin-internal ribosome entry site-enhanced green fluorescent protein (survivin-IRES-EGFP) in primary mouse marrow cells increased granulocyte macrophage-colony-forming units (CFU-GM) by 1.7- to 6.2-fold and the proportion of CFU-GM in S phase, compared to vector control. An antisense survivin construct decreased total and S-phase CFU-GM. These studies provide further evidence that survivin up-regulation by growth factors is not a consequence of cell cycle progression and strongly suggest that survivin is an important early event for cell cycle entry by CD34(+) cells.
凋亡抑制蛋白（IAP）survivin在增殖细胞如胎儿组织和癌症中表达。我们之前报道过survivin在正常成人CD34(+)细胞中表达且受生长因子调控。在此，我们检测了细胞周期进入前后CD34(+)细胞中survivin的表达，并证明了survivin在细胞周期调控和增殖中的作用。对已知人类IAPs的分析显示，在CD34(+)细胞中只有survivin受细胞因子调控。Survivin的表达与细胞周期进程一致。血小板生成素（Tpo）、Flt3配体（FL）和干细胞因子（SCF）对survivin的上调发生在视网膜母细胞瘤蛋白（Rb）低磷酸化（阳性）、Ki-67（阴性）和细胞周期蛋白D（阴性）的CD34(+)细胞中。定量实时逆转录-聚合酶链反应（RT-PCR）和多参数流式细胞术表明，Tpo、SCF和FL可增加静止G(0)期CD34(+)细胞中survivin的mRNA和蛋白水平，而不增加Ki-67的表达，这表明细胞因子刺激CD34(+)细胞中survivin的上调发生在G(0)期，即细胞进入G(1)期之前。PI3激酶/AKT和丝裂原活化蛋白激酶（MAPK(p42/44)）途径的选择性抑制在阻止细胞周期之前阻断了生长因子对survivin的上调。与载体对照相比，将survivin-内部核糖体进入位点-增强型绿色荧光蛋白（survivin-IRES-EGFP）逆转录病毒转导至原代小鼠骨髓细胞中，可使粒细胞巨噬细胞集落形成单位（CFU-GM）增加1.7至6.2倍，并增加S期CFU-GM的比例。反义survivin构建体可减少总CFU-GM和S期CFU-GM。这些研究进一步证明生长因子对survivin的上调不是细胞周期进程的结果，并强烈表明survivin是CD34(+)细胞进入细胞周期的一个重要早期事件。
