生物谷報道：RNA聚合酶II大亞基（Rpb1）是決定真核生物信使RNA（mRNA）轉(zhuǎn)錄起始和延伸的最主要的功能亞基，存在多種翻譯后修飾,。其中,，它的泛素化修飾和降解，不僅發(fā)生在DNA損傷引起的轉(zhuǎn)錄停滯過程中,，而且涉及到其他的轉(zhuǎn)錄障礙事件,。然而，在哺乳動物細(xì)胞中,，在非DNA損傷依賴過程中發(fā)揮作用的Rpb1的泛素連接酶還沒有被發(fā)現(xiàn),。健康科學(xué)研究所金穎研究員領(lǐng)導(dǎo)的分子發(fā)育生物學(xué)實驗室的最新研究表明Wwp2，小鼠的HECT家族的泛素E3連接酶,，是Rpb1新的泛素連結(jié)酶,。研究結(jié)果顯示在體內(nèi)和體外，小鼠的Wwp2均能特異性結(jié)合Rpb1并催化它的泛素化修飾,。有趣的是,，Wwp2 對Rpb1的結(jié)合和泛素化修飾既不依賴于Rpb1的磷酸化狀態(tài)也不依賴于DNA損傷。然而Wwp2的酶活性對于Rpb1的泛素化修飾是必需的,。進(jìn)一步的研究還顯示,，Wwp2和Rpb1的相互作用是通過Wwp2 的WW 結(jié)構(gòu)域和Rpb1的C端結(jié)構(gòu)域CTD介導(dǎo)的。當(dāng)下調(diào)Wwp2的表達(dá)水平時,，Rpb1的泛素化水平也隨之降低,，而蛋白水平則明顯升高。通過質(zhì)譜分析,，Rpb1 CTD結(jié)構(gòu)域中的6個賴氨酸殘基被確認(rèn)為Wwp2介導(dǎo)的泛素化位點,。這些結(jié)果顯示W(wǎng)wp2在Rpb1正常生理狀態(tài)下的表達(dá)調(diào)節(jié)中發(fā)揮重要功能。
該研究成果發(fā)表在07年8月的《分子細(xì)胞生物學(xué)》雜志上,。（援引健康科學(xué)研究所）
原始出處:
Molecular and Cellular Biology, August 2007, p. 5296-5305, Vol. 27, No. 15
0270-7306/07/$08.00+0 doi:10.1128/MCB.01667-06
Wwp2-Mediated Ubiquitination of the RNA Polymerase II Large Subunit in Mouse Embryonic Pluripotent Stem Cells
Hui Li,1,2,5 Zhihong Zhang,1 Beibei Wang,1 Junmei Zhang,3 Yingming Zhao,3 and Ying Jin1,2,4*
Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao Tong University School of Medicine, 225 South Chongqing Road, Shanghai 200025, China,1 Laboratory of Molecular Developmental Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China,2 Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390,3 Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai Jiao Tong University School of Medicine, Shanghai, China,4 Graduate School of Chinese Academy of Sciences, Beijing, China5
Received 6 September 2006/ Returned for modification 15 November 2006/ Accepted 13 May 2007
Ubiquitination and the degradation of the large subunit of RNA polymerase II, Rpb1, is not only involved in DNA damage-induced arrest but also in other transcription-obstructing events. However, the ubiquitin ligases responsible for DNA damage-independent processes in mammalian cells remain to be identified. Here, we identified Wwp2, a mouse HECT domain ubiquitin E3 ligase, as a novel ubiquitin ligase of Rpb1. We found that Wwp2 specifically interacted with mouse Rpb1 and targeted it for ubiquitination both in vitro and in vivo. Interestingly, the interaction with and ubiquitination of Rpb1 was dependent neither on its phosphorylation state nor on DNA damage. However, the enzymatic activity of Wwp2 was absolutely required for its ubiquitin modification of Rpb1. Furthermore, our study indicates that the interaction between Wwp2 and Rpb1 was mediated through WW domain of Wwp2 and C-terminal domain of Rpb1, respectively. Strikingly, downregulation of Wwp2 expression compromised Rpb1 ubiquitination and elevated its intracellular steady-state protein level significantly. Importantly, we identified six lysine residues in the C-terminal domain of Rpb1 as ubiquitin acceptor sites mediated by Wwp2. These results indicate that Wwp2 plays an important role in regulating expression of Rpb1 in normal physiological conditions.
