Arrest of cell cycle by Amida which is phosphorylated by Cdc2 kinase
Arrest of cell cycle by Amida which is phosphorylated by Cdc2 kinase
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DOI：
10.1023/a:1023479122934
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发表时间：
2003-04-01
影响因子：
4.3
通讯作者：
Miki, N
中科院分区：
文献类型：
--
作者：
Gan, YH;Taira, E;Miki, N
Amida是从大鼠海马cDNA文库中首次分离到的一种Arc相关蛋白。尽管先前的研究表明Amida mRNA在大鼠睾丸中主要表达并受发育调控，并且过表达诱导细胞凋亡，但Amida的功能仍不清楚。在这项研究中，我们发现过表达的Amida抑制细胞生长。流式细胞仪分析表明，Amida可使细胞周期阻滞在S期，并阻止细胞进入有丝分裂期。为了阐明Amida对细胞周期的影响，我们发现Amida在有丝分裂中与Cdc 2相互作用，并且Amida的过表达导致Cdc 2激酶活性的降低。DNA亲和柱层析显示Amida具有DNA结合能力。发现两个核定位信号之间的一个区域（aa，76-189）负责细胞生长抑制和DNA结合活性，暗示DNA结合活性可能是Amida抑制细胞周期所必需的。此外，Amida在体外被Cdc 2激酶磷酸化，并被鉴定为Amida的Ser-180磷酸化位点。AmidaS 180 G（消除Ser-180的磷酸化）具有更强的DNA结合活性。综上所述，这些数据表明，Amida可能在细胞周期中发挥重要作用，并可能部分受到Cdc 2激酶的调节。
Amida was first isolated from a rat hippocampal cDNA library as an Arc-associated protein. Although previous studies have shown that Amida mRNA is predominantly expressed and developmentally regulated in rat testis and overexpression induces apoptosis, the function of Amida remains unclear. In this study, we found that overexpression of Amida inhibited cell growth. Flow cytometry analysis showed that Amida caused cell cycle inhibition in the S-phase and blocked cell cycle from entry into mitosis. Attempting to elucidate Amida effect on the cell cycle, we found that Amida was interacted with Cdc2 in mitosis and Amida's overexpression resulted in a decrease in Cdc2 kinase activity. In addition, Amida showed DNA-binding ability with DNA-affinity column chromatography. A region (aa, 76-189) between the two nuclear localization signals was found to be responsible for cell growth inhibition and DNA-binding activity, implying that DNA-binding activity may be necessary for Amida to repress cell cycle. Moreover, Amida was phosphorylated by Cdc2 kinase in vitro and Ser-180 of Amida was identified as the phosphorylation site. Furthermore, AmidaS180G ( eliminate phosphorylation of Ser-180) showed stronger DNA-binding activity. Taken together, the data suggest that Amida may play an important role in cell cycle and may be partly regulated by Cdc2 kinase.