Department of Chemistry, University of California Riverside , Riverside, California 92521-0403, United States.
J Am Chem Soc. 2017 Sep 13;139(36):12426-12429. doi: 10.1021/jacs.7b07563. Epub 2017 Sep 5.
The guanine quadruplex (G4) structure in DNA is a secondary structure motif that plays important roles in DNA replication, transcriptional regulation, and maintenance of genomic stability. Here, we employed a quantitative mass spectrometry-based approach to profile the interaction proteomes of three well-defined G4 structures derived from the human telomere and the promoters of cMYC and cKIT genes. We identified SLIRP as a novel G4-interacting protein. We also demonstrated that the protein could bind directly with G4 DNA with K values in the low nanomolar range and revealed that the robust binding of the protein toward G4 DNA requires its RRM domain. We further assessed, by using CRISPR-Cas9-introduced affinity tag and ChIP-Seq analysis, the genome-wide occupancy of SLIRP, and showed that the protein binds preferentially to G-rich DNA sequences that can fold into G4 structures. Together, our results uncovered a novel cellular protein that can interact directly with G4 DNA, which underscored the complex regulatory networks involved in G4 biology.
DNA 中的鸟嘌呤四链体 (G4) 结构是一种二级结构基序，在 DNA 复制、转录调控和基因组稳定性维持中发挥重要作用。在这里，我们采用了一种基于定量质谱的方法来描绘三种明确的 G4 结构的相互作用蛋白质组，这些 G4 结构来源于人类端粒和 cMYC 和 cKIT 基因的启动子。我们鉴定出 SLIRP 是一种新的 G4 相互作用蛋白。我们还证明，该蛋白可以与 G4 DNA 直接结合，Kd 值在纳摩尔范围内，并揭示了该蛋白对 G4 DNA 的强结合需要其 RRM 结构域。我们进一步通过使用 CRISPR-Cas9 引入的亲和标签和 ChIP-Seq 分析，评估了 SLIRP 的全基因组占有率，并表明该蛋白优先结合可以折叠成 G4 结构的富含 G 的 DNA 序列。总之，我们的研究结果揭示了一种可以直接与 G4 DNA 相互作用的新型细胞蛋白，这突显了 G4 生物学中涉及的复杂调控网络。
