Valieva Maria E, Gerasimova Nadezhda S, Kudryashova Kseniya S, Kozlova Anastasia L, Kirpichnikov Mikhail P, Hu Qi, Botuyan Maria Victoria, Mer Georges, Feofanov Alexey V, Studitsky Vasily M
Biology Faculty, Lomonosov Moscow State University, Leninskie Gory 1, Moscow 119992, Russia.
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of Russian Academy of Sciences, Moscow 117997, Russia.
Cancers (Basel). 2017 Jan 6;9(1):3. doi: 10.3390/cancers9010003.
A correct chromatin structure is important for cell viability and is tightly regulated by numerous factors. Human protein complex FACT (facilitates chromatin transcription) is an essential factor involved in chromatin transcription and cancer development. Here FACT-dependent changes in the structure of single nucleosomes were studied with single-particle Förster resonance energy transfer (spFRET) microscopy using nucleosomes labeled with a donor-acceptor pair of fluorophores, which were attached to the adjacent gyres of DNA near the contact between H2A-H2B dimers. Human FACT and its version without the C-terminal domain (CTD) and the high mobility group (HMG) domain of the structure-specific recognition protein 1 (SSRP1) subunit did not change the structure of the nucleosomes, while FACT without the acidic C-terminal domains of the suppressor of Ty 16 (Spt16) and the SSRP1 subunits caused nucleosome aggregation. Proteolytic removal of histone tails significantly disturbed the nucleosome structure, inducing partial unwrapping of nucleosomal DNA. Human FACT reduced DNA unwrapping and stabilized the structure of tailless nucleosomes. CTD and/or HMG domains of SSRP1 are required for this FACT activity. In contrast, previously it has been shown that yeast FACT unfolds (reorganizes) nucleosomes using the CTD domain of SSRP1-like Pol I-binding protein 3 subunit (Pob3). Thus, yeast and human FACT complexes likely utilize the same domains for nucleosome reorganization and stabilization, respectively, and these processes are mechanistically similar.
正确的染色质结构对细胞活力至关重要，并受到众多因素的严格调控。人类蛋白质复合物FACT（促进染色质转录）是参与染色质转录和癌症发展的关键因素。在此，我们使用单颗粒Förster共振能量转移（spFRET）显微镜，对单核小体结构中依赖FACT的变化进行了研究，该显微镜使用了由荧光供体-受体对标记的核小体，这些荧光基团连接在H2A-H2B二聚体接触点附近DNA的相邻螺旋上。人类FACT及其缺少结构特异性识别蛋白1（SSRP1）亚基的C末端结构域（CTD）和高迁移率族（HMG）结构域的变体并未改变核小体的结构，而缺少Ty 16抑制因子（Spt16）和SSRP1亚基酸性C末端结构域的FACT则导致核小体聚集。蛋白酶解去除组蛋白尾巴会显著扰乱核小体结构，诱导核小体DNA部分解旋。人类FACT可减少DNA解旋并稳定无尾核小体的结构。SSRP1的CTD和/或HMG结构域是FACT发挥此活性所必需的。相比之下，此前研究表明酵母FACT利用类似SSRP1的RNA聚合酶I结合蛋白3亚基（Pob3）的CTD结构域使核小体解折叠（重新组织）。因此，酵母和人类FACT复合物可能分别利用相同的结构域进行核小体重组和稳定，并且这些过程在机制上是相似的。
