Architecture and ssDNA interaction of the Timeless-Tipin-RPA complex

Witosch, Justine; Wolf, Eva; Mizuno, Naoko

doi:10.1093/nar/gku960

Cited by 27 publications

(27 citation statements)

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“…As a core component of the replisome, Timeless is intimately associated with DNA synthesis at the replication fork (Yeeles et al, 2017). In vitro data show that the Timeless-Tipin complex can bind to ssDNA through RPA (Witosch et al, 2014), and the Swi1-Swi3 complex, the fission yeast orthologue of Timeless-Tipin, was also shown to bind DNA (Tanaka et al, 2010). Inspection of the amino acid sequence of human Timeless revealed the presence of a conserved domain in its C-terminal half (residues 816-954; Fig 2A), with a predicted fold similarity to the myb-like proteins of the homeodomain-like superfamily, that bind double-stranded DNA (dsDNA) with a tandem repeat of 3-helix bundles (named here N-term and C-term).…”

Section: Identification and Characterisation Of A Timeless Dna-bindinmentioning

confidence: 99%

“…FPC components associate with the replication fork via direct interactions with the CMG replicative helicase and replicative polymerases a, d and e (Nedelcheva et al, 2005;Numata et al, 2010;Cho et al, 2013;Bastia et al, 2016;Kilkenny et al, 2017;Bareti c et al, 2020). They also interact with DNA both directly (Tanaka et al, 2010) and indirectly via replication protein A (RPA) (Witosch et al, 2014). These interactions allow the FPC to remain at the fork, which ensures a normal speed of DNA synthesis (Yeeles et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

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Timeless couples G‐quadruplex detection with processing by DDX 11 helicase during DNA replication

et al. 2020

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Regions of the genome with the potential to form secondary DNA structures pose a frequent and significant impediment to DNA replication and must be actively managed in order to preserve genetic and epigenetic integrity. How the replisome detects and responds to secondary structures is poorly understood. Here, we show that a core component of the fork protection complex in the eukaryotic replisome, Timeless, harbours in its C‐terminal region a previously unappreciated DNA‐binding domain that exhibits specific binding to G‐quadruplex (G4) DNA structures. We show that this domain contributes to maintaining processive replication through G4‐forming sequences, and exhibits partial redundancy with an adjacent PARP‐binding domain. Further, this function of Timeless requires interaction with and activity of the helicase DDX11. Loss of both Timeless and DDX11 causes epigenetic instability at G4‐forming sequences and DNA damage. Our findings indicate that Timeless contributes to the ability of the replisome to sense replication‐hindering G4 formation and ensures the prompt resolution of these structures by DDX11 to maintain processive DNA synthesis.

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Section: Identification and Characterisation Of A Timeless Dna-bindinmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Timeless couples G‐quadruplex detection with processing by DDX 11 helicase during DNA replication

et al. 2020

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“…FPC components associate with the replication fork via direct interactions with the CMG replicative helicase and replicative polymerases α, δ and ε [16][17][18][19] . They also interact with DNA both directly 20 and indirectly via replication protein A 21 . These interactions allow the FPC to remain at the fork, which ensures a normal speed of DNA synthesis 22 .…”

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confidence: 99%

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

Lerner

Holzer

Kilkenny

et al. 2019

Preprint

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& 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...

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“…As the first 3D structural analysis of a TIMELESS protein, the cryo electron microscopy structure and biochemical studies of a complex of mTIM with its interaction partner Tipin (Timeless interacting protein) and the replication protein A (RPA) has recently been reported [148]. This work has shed light on the 3D architecture of the Timeless-Tipin-RPA complex and the regulation of the recruitment of the mTIM-Tipin complex to the DNA replication fork via RPA conformational changes.…”

Section: A Final Note On Mammalian Timeless: a Link Between The Circamentioning

confidence: 99%

How is the inner circadian clock controlled by interactive clock proteins?

Merbitz-Zahradnik

Wolf

2015

FEBS Letters

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Edited by Wilhelm JustKeywords: Circadian rhythm 3D crystal structure Drosophila and mammalian clock protein Circadian clock mechanism a b s t r a c t Most internationally travelled researchers will have encountered jetlag. If not, working odd hours makes most of us feel somehow dysfunctional. How can all this be linked to circadian rhythms and circadian clocks? In this review, we define circadian clocks, their composition and underlying molecular mechanisms. We describe and discuss recent crystal structures of Drosophila and mammalian core clock components and the enormous impact they had on the understanding of circadian clock mechanisms. Finally, we highlight the importance of circadian clocks for the daily regulation of human/mammalian physiology and show connections to overall fitness, health and disease.

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Architecture and ssDNA interaction of the Timeless-Tipin-RPA complex

Cited by 27 publications

References 64 publications

Timeless couples G‐quadruplex detection with processing by DDX 11 helicase during DNA replication

Timeless couples G‐quadruplex detection with processing by DDX 11 helicase during DNA replication

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

How is the inner circadian clock controlled by interactive clock proteins?

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