-Actin regulates interleukin 6-induced p21 transcription by interacting with the Rpb5 and Rpb7 subunits of RNA polymerase II
-Actin regulates interleukin 6-induced p21 transcription by interacting with the Rpb5 and Rpb7 subunits of RNA polymerase II
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-肌动蛋白通过与 RNA 聚合酶 II 的 Rpb5 和 Rpb7 亚基相互作用来调节白细胞介素 6 诱导的 p21 转录
DOI：
10.1080/19768354.2016.1224204
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发表时间：
2016
影响因子：
2.9
通讯作者：
Mi Donghui
中科院分区：
文献类型：
--
作者：
Tian Xiujuan;Qi Wenjing;Chen Hongyu;Zeng Xianlu;Han Liping;Mi Donghui
在起始前复合物中，RNA解旋酶A与β-肌动蛋白相互作用，并作为连接核肌动蛋白与RNA聚合酶II （Pol II）的桥接因子。此外，β-肌动蛋白通过与正转录延伸因子Cdk9相互作用参与Pol ii依赖性转录延伸。然而，β-肌动蛋白和Pol II之间的许多关系仍有待确定。在白细胞介素6 （IL-6）诱导的p21表达模型中，我们证实β-actin敲低可降低p21的表达。免疫荧光分析显示，IL-6处理的细胞中β-actin和Pol II的共定位明显增加。已知Rpb5、Rpb6和Rpb7亚基位于酶的表面。接下来，我们构建了重组pcDNA-HA-Rpb5、pcDNA-HA-Rpb6和pcDNA-HA-Rpb7质粒，并在HepG2细胞中表达了三个聚合酶II亚基。我们发现HA-Rpb5和HA-Rpb7可以免疫沉淀β-肌动蛋白。谷胱甘肽s转移酶下拉实验显示β-肌动蛋白与Rpb5和rpb7在体外相关。此外，细胞中Rpb5和Rpb7的过表达显著降低了p21的表达，表明Rpb5和Rpb7与β-actin竞争性相互作用。这项研究表明，β-actin通过直接的蛋白质相互作用与Pol II亚基结合，并为Pol II转录调控提供了基本的见解。DAPI: 4′，6′-二氨基-2-苯基吲哚；胎牛血清；HAT：组蛋白乙酰转移酶；HDAC：组蛋白脱乙酰酶；销售税:Glutathione-S-transferase;IL-6：白细胞介素6；PICs：起始前配合物；Pol II: RNA聚合酶II；RHA: RNA解旋酶A；siRNA：小干扰RNA
In pre-initiation complexes, RNA helicase A interacts with β-actin and acts as a bridging factor linking nuclear actin with RNA polymerase II (Pol II). In addition, β-actin participates in Pol II-dependent transcription elongation by interacting with the positive transcription elongation factor Cdk9. However, many relationships between β-actin and Pol II remain to be identified. In an interleukin 6 (IL-6)-induced p21 expression model, we demonstrated that β-actin knockdown reduced p21 expression. Immunofluorescence analysis showed that the colocalization of β-actin and Pol II increased significantly in cells treated with IL-6. It is known that the Rpb5, Rpb6 and Rpb7 subunits are located at the surface of the enzyme. We next constructed recombinant pcDNA-HA-Rpb5, pcDNA-HA-Rpb6 and pcDNA-HA-Rpb7 plasmids and expressed the three polymerase II subunits in HepG2 cells. We found that β-actin could be immunoprecipitated with HA-Rpb5 and HA-Rpb7. A Glutathione-S-transferase pull-down assay revealed that β-actin was associated with Rpb5 and Rpb7in vitro. Furthermore, overexpression of Rpb5 and Rpb7 in cells reduced p21 expression significantly, suggesting that Rpb5 and Rpb7 competitively interact with β-actin. This study shows that β-actin associates with Pol II subunits through direct protein-protein interactions and provides fundamental insight into Pol II transcriptional regulation.Abbreviations: DAPI: 4′,6′-diamidino-2-phenylindole; FBS: fetal bovine serum; HAT: histone acetyltransferase; HDAC: histone deacetylase; GST: Glutathione-S-transferase; IL-6: interleukin 6; PICs: pre-initiation complexes; Pol II: RNA polymerase II; RHA: RNA helicase A; siRNA: small interfering RNA