Nuclear localization of Lyn tyrosine kinase mediated by inhibition of its kinase activity

Src-family kinases, cytoplasmic enzymes that participate in various signaling events, are found at not only the plasma membrane but also subcellular compartments, such as the nucleus, the Golgi apparatus and late endosomes/lysosomes. Lyn, a member of the Src-family kinases, is known to play a role in DNA damage response and cell cycle control in the nucleus. However, it is still unclear how the localization of Lyn to the nucleus is regulated. Here, we investigated the mechanism of the distribution of Lyn between the cytoplasm and the nucleus in epitheloid HeLa cells and hematopoietic THP-1 cells. Lyn was definitely detected in purified nuclei by immunofluorescence and immunoblotting analyses. Nuclear accumulation of Lyn was enhanced upon treatment of cells with leptomycin B (LMB), an inhibitor Of Crm1-mediated nuclear export. Moreover, Lyn mutants lacking the sites for lipid modification were highly accumulated in the nucleus upon LMB treatment. Intriguingly, inhibition of the kinase activity of Lyn by SU6656, Csk overexpression, or point mutation in the ATP-binding site induced an increase in nuclear Lyn levels. These results Suggest that Lyn being imported into and rapidly exported from the nucleus preferentially accumulates in the nucleus by inhibition of the kinase activity and lipid modification. (c) 2008 Elsevier Inc. All rights reserved.

Src家族激酶是参与多种信号传导事件的胞质酶，不仅存在于质膜，还存在于亚细胞区室，如细胞核、高尔基体以及晚期内体/溶酶体。Lyn是Src家族激酶的成员，已知其在细胞核的DNA损伤应答和细胞周期控制中发挥作用。然而，Lyn在细胞核的定位是如何被调控的仍不清楚。在此，我们研究了上皮样HeLa细胞和造血THP - 1细胞中Lyn在细胞质和细胞核之间分布的机制。通过免疫荧光和免疫印迹分析，在纯化的细胞核中明确检测到Lyn。用 leptomycin B（LMB，一种Crm1介导的核输出抑制剂）处理细胞后，Lyn在细胞核中的积累增强。此外，缺失脂质修饰位点的Lyn突变体在LMB处理后在细胞核中高度积累。有趣的是，用SU6656抑制Lyn的激酶活性、Csk过表达或ATP结合位点的点突变导致细胞核中Lyn水平升高。这些结果表明，Lyn优先通过激酶活性抑制和脂质修饰在细胞核中积累，它被输入细胞核并迅速从细胞核输出。（c）2008 Elsevier Inc.保留所有权利。