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Involvement of CK2 for the nuclear dynamic function during the progression of cell cycle.

CK2 在细胞周期进展过程中参与核动态功能。

基本信息

批准号：
20570137
负责人：
HOMMA MiwakoK.
金额：
$ 3.16万
依托单位：
Fukushima Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2008
资助国家：
日本
起止时间：
2008 至 2010
项目状态：
已结题

项目摘要

Protein kinase CK2 is a highly conserved serine/threonine kinase that functions in multiple cellular processes including cell growth and development. It is typically found in tetrameric complexes consisting of two catalytic α and/or α' subunits, and two regulatory β subunits. We reported the cell-cycle dependent association of CK2 with adenomatous polyposis coli (APC) that regulates CK2 activity, and identified its downstream target as eukaryotic translation initiation factor 5 (eIF5). Here we demonstrate that a pool of CK2α subunits translocate into the nucleus in G1 phase of the cell cycle, when serum-starved normal human fibroblasts are simulated by FBS, whereas the β subunits localize to the nuclear fraction later than α. Translocation of αβ subunits associate with enhanced phosphorylation of these molecules. Pretreatment with a potent ATP competitive inhibitors reduce nuclear localization of CK2α after FBS stimuli. Similarly, phosphorylation site mutants in CK2α partially inhibit their accumulation in the nuclear fraction following FBS stimuli. These results suggest the involvement of multiple kinases that phosphorylate CK2 for the growth factor-induced nuclear localization. Co-immunoprecipitation experiments using anti-CK2α antibodies, followed by mass spectrometry analysis, identified nuclear proteins that associate with CK2α in a cell -cycle dependent manner. As one of these proteins, hnRNPs is also a phosphoprotein in vivo. FRAP experiments on living stable cell lines expressing GFP-CK2α protein indicate that the nucleocytoplasmic distribution of CK2 subunits may be regulated in a cell cycle-dependent manner. These results support a role for CK2 in response to growth stimuli, and also the possible involvement of CK2α subunit as isolated entities exerting specific functions in complex with other cellular partners in the nuclear fraction, during the progression of cell cycle.

蛋白激酶CK2是一种高度保守的丝氨酸/苏氨酸激酶，参与多种细胞过程，包括细胞的生长和发育。它通常存在于由两个催化的α和/或α‘亚基和两个调节的β亚基组成的四聚体复合体中。我们报道了CK2与调节CK2活性的腺瘤性息肉病(APC)之间的细胞周期依赖性关联，并确定其下游靶点为真核细胞翻译起始因子5(EIF5)。在这里，我们证明了当血清饥饿的正常人成纤维细胞被胎牛血清模拟时，CK2α亚基在细胞周期的G1期移位到细胞核，而β亚基定位到细胞核部分的时间晚于α。αβ亚基的易位与这些分子的磷酸化增强有关。用有效的三磷酸腺苷竞争性抑制剂预处理可减少胎牛血清刺激后CK2α的核定位。类似地，CK2α中的磷酸化位点突变部分地抑制了它们在FBS刺激后在核部分的积聚。这些结果表明，在生长因子诱导的核定位中，参与了多个磷酸化CK2的激酶的参与。使用抗CK2α抗体的免疫共沉淀实验，随后进行质谱分析，鉴定了与CK2α相关的核蛋白，并以细胞周期依赖的方式。作为其中的一种蛋白质，hnRNPs在体内也是一种磷蛋白。在稳定表达GFP-CK2α蛋白的活细胞株上的FRAP实验表明，CK2亚基的核质分布可能以细胞周期依赖的方式调节。这些结果支持了CK2在对生长刺激的反应中的作用，也支持了CK2α亚基作为孤立的实体在细胞周期的进程中与核部分中的其他细胞伙伴一起发挥特定功能的可能性。