Dictyostelium differentiation-inducing factor-3 activates glycogen synthase kinase-3β and degrades cyclin D1 in mammalian cells
Dictyostelium differentiation-inducing factor-3 activates glycogen synthase kinase-3β and degrades cyclin D1 in mammalian cells
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DOI：
10.1074/jbc.m205768200
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发表时间：
2003-03-14
影响因子：
4.8
通讯作者：
Sasaguri, T
中科院分区：
文献类型：
--
作者：
Takahashi-Yanaga, F;Taba, Y;Sasaguri, T
为了寻找可用于治疗癌症和其他增殖性疾病的化学物质，我们主要使用HeLa细胞研究了哺乳动物细胞中的DIFs（Dictyosteoprotein-differentiation-inducing factors）的信号转导。尽管DIF-1和DIF-3都通过诱导G（0）/G（1）停滞来强烈抑制细胞增殖，但DIF-3比DIF-1更有效。DIF-3抑制细胞周期蛋白D1的表达在mRNA和蛋白质水平，而细胞周期蛋白D1的过度表达推翻DIF-3诱导的细胞周期阻滞。DIF-3诱导的细胞周期蛋白D1蛋白量的减少先于细胞周期蛋白D1 mRNA水平的减少。细胞周期蛋白D1蛋白的减少似乎是由加速的蛋白水解引起的，因为它被蛋白酶体抑制剂N-乙酰基-Leu-Leu-norleucinal废除。DIF-3诱导的细胞周期蛋白D1降解也被氯化锂（糖原合成酶激酶-3 β（GSK-3 β）的抑制剂）阻止，这表明DIF-3通过激活GSK-3 β诱导细胞周期蛋白D1蛋白水解。事实上，DIF-3使GSK-3 β上的Ser（9）去磷酸化并使酪氨酸磷酸化，并且在体外激酶测定中它刺激GSK-3 β活性。此外，通过免疫荧光显微镜和亚细胞蛋白组分的免疫印迹，显示DIF-3诱导GSK-3 β的核转位。这些结果表明，DIF-3激活GSK-3 β以加速细胞周期蛋白D1的蛋白水解，并且该机制参与了DIF-3诱导的哺乳动物细胞G（0）/G（1）阻滞。
In search of chemical substances applicable for the treatment of cancer and other proliferative disorders, we studied the signal transduction of Dictyostelium differentiation-inducing factors (DIFs) in mammalian cells mainly using HeLa cells. Although DIF-1 and DIF-3 both strongly inhibited cell proliferation by inducing G(0)/G(1) arrest, DIF-3 was more effective than DIF-1. DIF-3 suppressed cyclin D1 expression at both mRNA and protein levels, whereas the overexpression of cyclin D1 overrode DIF-3-induced cell cycle arrest. The DIF-3-induced decrease in the amount of cyclin D1 protein preceded the reduction in the level of cyclin D1 mRNA. The decrease in cyclin D1 protein seemed to be caused by accelerated proteolysis, since it was abrogated by N-acetyl-Leu-Leu-norleucinal, a proteasome inhibitor. DIF-3-induced degradation of cyclin D1 was also prevented by treatment with lithium chloride, an inhibitor of glycogen synthase kinase-3beta (GSK-3beta), suggesting that DIF-3 induced cyclin D1 proteolysis through the activation of GSK-3beta. Indeed, DIF-3 dephosphorylated Ser(9) and phosphorylated tyrosine on GSK-3beta, and it stimulated GSK-3beta activity in an in vitro kinase assay. Moreover, DIF-3 was revealed to induce the nuclear translocation of GSK-3beta by immunofluorescent microscopy and immunoblotting of subcellular protein fractions. These results suggested that DIF-3 activates GSK-3beta to accelerate the proteolysis of cyclin D1 and that this mechanism is involved in the DIF-3-induced G(0)/G(1) arrest in mammalian cells.