Timeless couples G quadruplex detection with processing by DDX11 during DNA replication

Abstract

& equal contribution2 Regions of the genome with the potential to form secondary structure pose a frequent and significant impediment to DNA replication and must be actively managed in order to preserve genetic and epigenetic integrity. The fork protection complex (FPC), a conserved group of replisome-associated proteins including Timeless, Tipin, and Claspin, plays an important role in maintaining efficient replisome activation, ensuring optimum fork rates, sister chromatid cohesion and checkpoint function. It also helps maintain the stability of sequences prone to secondary structure formation through an incompletely understood mechanism. Here, we report a previously unappreciated DNA binding domain in the C-terminus of Timeless, which exhibits specific binding to G quadruplex (G4) structures. We show that, in vivo, both the C-terminus of Timeless and the DDX11 helicase act collaboratively to ensure processive replication of G4 structures to prevent genetic and epigenetic instability.DNA can create significant impediments to its own replication through formation of secondary structures. When unwound, certain sequences, often repetitive or of low complexity, can adopt a variety of non-B form structures, including hairpins, cruciforms, triplexes and quadruplexes 1 . It is becoming clear that secondary structure formation is a frequent event during replication, even at genomically abundant sequences previously thought not to be a major source of difficulty 2 . To prevent such sequences causing havoc with the genetic and epigenetic stability of the genome, cells deploy an intricate network of activities to counteract secondary structure formation and limit its effects. These activities include proteins that bind and destabilise DNA structures and specialised helicases that unwind them 3 . In addition, the repriming activity of PrimPol can be deployed to confine a structure into a minimal region of single stranded DNA, limiting the potential dangers of exposing extensive ssDNA in a stalled replisome 2,4 . G quadruplexes (G4s) are one of the most intensively studied and potent structural replication impediments. G4s arise in consequence of the ability of guanine to form Hoogsteen base-paired quartets 5 . In favourable sequence contexts, comprising runs of dG separated by variable numbers of non-G bases, stacks of G quartets form G4s secondary structures. Current estimates suggest that over 700,000 sites in the human genome have the potential to form G4s 6 . While some of these G4s may have important roles in genome physiology, all pose a potential threat to DNA replication and sites with G4-forming potential have been linked to both genetic and epigenetic instability 7,8 .3Precisely how DNA structures are detected and resolved by the replication machinery remains unclear. Many of the factors involved in processing G4 secondary structures, for instance FANCJ and REV1 9-13 , do not appear to be constitutive components of the replisome 14 . It is thus likely that core components of the replisome will act as 'first respo...

Mean field theory of jamming of nonspherical particles

“…It would be interesting to extend the theory in Ref. [58] to the present case to study how the rheology of nonspherical particles differs from that of spherical ones.…”

“…An interesting question is how this difference affects the steady-state rheology near the jamming transition point. For spherical particles, it has been shown that the minimal eigenfrequency of D(ω) controls the viscosity near the jamming transition point [58]. It would be interesting to extend the theory in Ref.…”

A Timeless Tale: G4 structure recognition by the fork protection complex triggers unwinding by DDX 11 helicase

“…G4 motifs are quite common in genomes, with over 700,000 sites in the human genome. A new study by the Sale and Pellegrini laboratories (Lerner et al , ) has provided a significant step forward in understanding how the replisome senses G4 DNA in order to facilitate G4 unwinding and replication.…”

“…Now, the new data from Lerner et al () shed light on how Timeless/Tof1 can facilitate replication through DNA structures. The authors identify a myb‐like DNA‐binding domain in the C‐terminus of Timeless that preferentially binds to G4 DNA with ten‐times higher affinity than to duplex DNA.…”

“…The latter was shown using an in vivo genetic assay that can detect replication efficiency of a G4 motif near the promoter of the chicken Bu‐1 gene. Prolonged pausing of leading strand replication at this motif leads to loss of epigenetic information around the promoter of the gene, and a permanent and heritable change in its expression that can be easily detected (Lerner et al , ). The DDX11 helicase was previously shown to interact with Timeless, and this interaction enhanced its DNA unwinding activity, including G4 unwinding (Calì et al , ).…”