近日,，國際雜志Nucleic Acids Research刊登了北京生命科學(xué)研究所葉克窮實(shí)驗(yàn)室和首都師范大學(xué)許興智實(shí)驗(yàn)室合作的最新研究成果“Structural mechanism of the phosphorylation-dependent dimerization of the MDC1 forkhead-associated domain,，”文章中，作者揭示了哺乳動物修復(fù)因子MDC1自身二聚化調(diào)節(jié)新的分子機(jī)制,。
編碼基因的DNA分子時刻會受各種因子(如紫外線,、自由基)的破壞發(fā)生損傷，而細(xì)胞能監(jiān)視和修復(fù)各種DNA損傷,。DNA雙鏈斷裂是最嚴(yán)重的損傷,，在斷裂發(fā)生后，ATM蛋白激酶得到激活,，磷酸化一系列底物,，包括斷裂位點(diǎn)附近的組蛋白H2AX。MDC1是哺乳動物修復(fù)DNA雙鏈斷裂的重要因子,，它通過其羰端BRCT結(jié)構(gòu)域識別磷酸化的H2AX,，從而定位在斷裂位點(diǎn)，它同時招募多個修復(fù)因子參與修復(fù)過程,。MDC1氮端含有一個能結(jié)合磷酸化蘇氨酸的FHA結(jié)構(gòu)域,，但其功能仍不清楚。
該論文通過蛋白晶體學(xué),、生物化學(xué)和細(xì)胞生物學(xué)等多種手段發(fā)現(xiàn),，在DNA損傷發(fā)生后，F(xiàn)HA結(jié)構(gòu)域介導(dǎo)依賴于MDC1磷酸化的二聚化,。作者發(fā)現(xiàn)MDC1 FHA結(jié)構(gòu)域自身能形成不太穩(wěn)定的二聚體,。在DNA損傷后，ATM激酶磷酸化MDC1第4位的蘇氨酸,，然后FHA結(jié)構(gòu)域反式結(jié)合另一個MDC1分子上磷酸化的第4位蘇氨酸,，從而形成穩(wěn)定的二聚體。作者還通過細(xì)胞學(xué)實(shí)驗(yàn)發(fā)現(xiàn),，MDC1二聚化能促進(jìn)其定位于DNA損傷位點(diǎn),。該工作揭示了MDC1 功能受其自身二聚化調(diào)節(jié)的新的分子機(jī)制。
首都師范大學(xué)的劉金平和我所研究生羅樹坤為本文的共同第一作者,。葉克窮博士和首都師范大學(xué)的許興智教授為本文的通訊作者,。在北京生命科學(xué)研究所的研究工作受到科技部和北京市政府的支持。（生物谷Bioon.com）
doi:10.1093/nar/gkr1296
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Structural mechanism of the phosphorylation-dependent dimerization of the MDC1 forkhead-associated domain
Jinping Liu1, Shukun Luo2,3, Hongchang Zhao1, Ji Liao1, Jing Li1, Chunying Yang4, Bo Xu4, David F. Stern5, Xingzhi Xu1,* and Keqiong Ye3,*
MDC1 is a key mediator of the DNA-damage response in mammals with several phosphorylation-dependent protein interaction domains. The function of its N-terminal forkhead-associated (FHA) domain remains elusive. Here, we show with structural, biochemical and cellular data that the FHA domain mediates phosphorylation-dependent dimerization of MDC1 in response to DNA damage. Crystal structures of the FHA domain reveal a face-to-face dimer with pseudo-dyad symmetry. We found that the FHA domain recognizes phosphothreonine 4 (pT4) at the N-terminus of MDC1 and determined its crystal structure in complex with a pT4 peptide. Biochemical analysis further revealed that in the dimer, the FHA domain binds in trans to pT4 from the other subunit, which greatly stabilizes the otherwise unstable dimer. We show that T4 is phosphorylated primarily by ATM upon DNA damage. MDC1 mutants with the FHA domain deleted or impaired in its ability to dimerize formed fewer foci at DNA-damage sites, but the localization defect was largely rescued by an artificial dimerization module, suggesting that dimerization is the primary function of the MDC1 FHA domain. Our results suggest a novel mechanism for the regulation of MDC1 function through T4 phosphorylation and FHA-mediated dimerization.
