Abstract
The Tof1–Csm3 fork protection complex has a central role in the replisome—it promotes the progression of DNA replication forks and protects them when they stall, while also enabling cohesion establishment and checkpoint responses. Here, I present the crystal structure of the Tof1–Csm3 complex from Chaetomium thermophilum at 3.1 Å resolution. The structure reveals that both proteins together form an extended alpha helical repeat structure, which suggests a mechanical or scaffolding role for the complex. Expanding on this idea, I characterize a DNA interacting region and a cancer-associated Mrc1 binding site. This study provides the molecular basis for understanding the functions of the Tof1–Csm3 complex, its human orthologue the Timeless–Tipin complex and additionally the Drosophila circadian rhythm protein Timeless.
Highlights
As well as error-free DNA synthesis, the eukaryotic replication fork must coordinate processes such as establishing chromosome cohesion, activating the S phase checkpoint and transferring epigenetic material
The coordination of all of these processes is required to maintain genome stability [1]. This becomes especially important in conditions of replication stress, which is a common feature of cancer cells [2,3]
To determine the structure of the Tof1–Csm3 complex, multiple constructs from the mildly thermophilic fungus Chaetomium thermophilum were screened for purification and expression
Summary
As well as error-free DNA synthesis, the eukaryotic replication fork must coordinate processes such as establishing chromosome cohesion, activating the S phase checkpoint and transferring epigenetic material. The Tof1–Csm fork protection complex was identified as a non-essential chromosome cohesion factor [7,8] that interacts with Topoisomerase 1 [9] to link concatenation and fork regulation [10]. It has an important role in protecting stalled forks and enabling their restart [11,12,13], coupling the replicative helicases and polymerases [14], promoting fork progression [15,16], mediating the S phase checkpoint [17,18] and maintaining genome stability at CAG repeats [19]. It has been suggested that mammalian Timeless links the circadian rhythm with DNA replication [32,33]
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