DNA-dependent protein kinase (DNA-PK) phosphorylates nuclear DNA helicase II/RNA helicase A and hnRNP proteins in an RNA-dependent manner
DNA-dependent protein kinase (DNA-PK) phosphorylates nuclear DNA helicase II/RNA helicase A and hnRNP proteins in an RNA-dependent manner
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DOI：
10.1093/nar/gkg933
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发表时间：
2004-01-01
影响因子：
14.9
通讯作者：
Grosse, F
中科院分区：
文献类型：
--
作者：
Zhang, SS;Schlott, B;Grosse, F
通过免疫沉淀和凝胶过滤层析，在 HeLa 细胞的核提取物中发现了 Ku 抗原与核 DNA 解旋酶 II (NDH II)（也称为 RNA 解旋酶 A (RHA)）的 RNA 依赖性关联。 Ku 抗原和 NDH II 均与 hnRNP 复合物相关。二维凝胶电泳显示，Ku 抗原与 hnRNP C、K、J、H 和 F 相关性最高，但与其他蛋白（例如 hnRNP A1）显然不相关。出乎意料的是，包含Ku抗原作为DNA结合亚基的DNA依赖性蛋白激酶（DNA-PK）以RNA依赖性方式磷酸化hnRNP蛋白。 DNA-PK 还在 RNA 存在的情况下磷酸化重组 NDH II。 RNA 结合测定显示 DNA-PK 对聚 (rG) 具有偏好，但对聚 (rA)、聚 (rC) 或聚 (rU) 不具有偏好。 DNA-PK 的这种 RNA 结合亲和力可归因于其 Ku86 亚基。一致地，poly(rG) 最强烈地刺激 DNA-PK 催化的 NDH II 磷酸化。 RNA干扰研究表明，NDH II的表达抑制改变了hnRNP C的核分布，而沉默DNA-PK则改变了NDH II和hnRNP C的亚核分布。这些结果支持这样的观点，即DNA-PK还可以作为RNA依赖性蛋白激酶来调节RNA代谢的某些方面，例如RNA加工和运输。
An RNA-dependent association of Ku antigen with nuclear DNA helicase II (NDH II), alternatively named RNA helicase A (RHA), was found in nuclear extracts of HeLa cells by immunoprecipitation and by gel filtration chromatography. Both Ku antigen and NDH II were associated with hnRNP complexes. Two-dimensional gel electrophoresis showed that Ku antigen was most abundantly associated with hnRNP C, K, J, H and F, but apparently not with others, such as hnRNP A1. Unexpectedly, DNA-dependent protein kinase (DNA-PK), which comprises Ku antigen as the DNA binding subunit, phosphorylated hnRNP proteins in an RNA-dependent manner. DNA-PK also phosphorylated recombinant NDH II in the presence of RNA. RNA binding assays displayed a preference of DNA-PK for poly(rG), but not for poly(rA), poly(rC) or poly(rU). This RNA binding affinity of DNA-PK can be ascribed to its Ku86 subunit. Consistently, poly(rG) most strongly stimulated the DNA-PK-catalyzed phosphorylation of NDH II. RNA interference studies revealed that a suppressed expression of NDH II altered the nuclear distribution of hnRNP C, while silencing DNA-PK changed the subnuclear distribution of NDH II and hnRNP C. These results support the view that DNA-PK can also function as an RNA-dependent protein kinase to regulate some aspects of RNA metabolism, such as RNA processing and transport.