Timeless collaborates with DDX11 to ensure processive replication of G4-forming DNA [RNA-seq]

Regions of DNA with the potential to form secondary structure pose a frequent and significant impediment to DNA replication that must be actively countered by cells in order to preserve genetic and epigenetic integrity. The fork protection complex (FPC), a conserved group of replisome-associated proteins, comprising Timeless, plays an important role in maintaining efficient replisome function. A previously unappreciated DNA binding domain in the C terminus of Timeless, which exhibits specificity towards G4 structures, acts in collaboration with the DDX11 helicase to ensure replication of G4 structures in vivo and maintenance of genetic and epigenetic stability. Using RNA-seq experiments, we show that (i) Timeless and DDX11 share common gene targets, (ii) the promoter of Timeless- and DDX11-dependent genes are enriched in G4 structures in the vicinity of their promoter and (iii) both Timeless and DDX11 share a common set of gene targets with the FANCJ helicase known to maintain epigenetic stability in the vicinity of G4 structures. RNA-seq data of WT, TIMELESS, DDX11 and FANCJ KO DT40 cells

具备形成二级结构潜能的DNA区域，常会对DNA复制造成频繁且显著的阻碍；细胞必须主动抵御此类阻碍，方能维持遗传与表观遗传的完整性。叉保护复合体（fork protection complex, FPC）是一类保守的复制体相关蛋白复合体，其组分包含Timeless蛋白，在维持复制体高效运作中发挥关键作用。此前尚未被认知的Timeless蛋白C端DNA结合结构域，对G-四链体（G4）结构具有特异性结合活性，可与DDX11解旋酶协同作用，保障G-四链体结构在体内的顺利复制，并维持遗传与表观遗传稳定性。本研究通过RNA测序（RNA-seq）实验证实：其一，Timeless与DDX11共享共同的基因靶点；其二，依赖Timeless与DDX11的基因，其启动子区域附近富集G-四链体结构；其三，Timeless与DDX11均与FANCJ解旋酶共享一套共同的基因靶点，而FANCJ解旋酶已知可在G-四链体结构附近维持表观遗传稳定性。野生型（WT）、Timeless基因敲除（KO）、DDX11基因敲除及FANCJ基因敲除的DT40细胞的RNA测序数据