The human CTF4-orthologue AND-1 interacts with DNA polymerase α/primase via its unique C-terminal HMG box.

A dynamic multi-protein assembly known as the replisome is responsible for DNA synthesis in eukaryotic cells. In yeast, the hub protein Ctf4 bridges DNA helicase and DNA polymerase and recruits factors with roles in metabolic processes coupled to DNA replication. An important question in DNA replication is the extent to which the molecular architecture of the replisome is conserved between yeast and higher eukaryotes.

Targeting the Genome-Stability Hub Ctf4 by Stapled-Peptide Design.

The exploitation of synthetic lethality by small-molecule targeting of pathways that maintain genomic stability is an attractive chemotherapeutic approach. The Ctf4/AND-1 protein hub, which links DNA replication, repair, and chromosome segregation, represents a novel target for the synthetic lethality approach. Herein, we report the design, optimization, and validation of double-click stapled peptides encoding the Ctf4-interacting peptide (CIP) of the replicative helicase subunit Sld5.

Ctf4 Is a Hub in the Eukaryotic Replisome that Links Multiple CIP-Box Proteins to the CMG Helicase.

Replisome assembly at eukaryotic replication forks connects the DNA helicase to DNA polymerases and many other factors. The helicase binds the leading-strand polymerase directly, but is connected to the Pol α lagging-strand polymerase by the trimeric adaptor Ctf4. Here, we identify new Ctf4 partners in addition to Pol α and helicase, all of which contain a "Ctf4-interacting-peptide" or CIP-box.

Structure of human Cdc45 and implications for CMG helicase function.

Cell division cycle protein 45 (Cdc45) is required for DNA synthesis during genome duplication, as a component of the Cdc45-MCM-GINS (CMG) helicase. Despite its essential biological function, its biochemical role in DNA replication has remained elusive. Here we report the 2.

Solution structure of the SGTA dimerisation domain and investigation of its interactions with the ubiquitin-like domains of BAG6 and UBL4A.

Background: The BAG6 complex resides in the cytosol and acts as a sorting point to target diverse hydrophobic protein substrates along their appropriate paths, including proteasomal degradation and ER membrane insertion. Composed of a trimeric complex of BAG6, TRC35 and UBL4A, the BAG6 complex is closely associated with SGTA, a co-chaperone from which it can obtain hydrophobic substrates.

Methodology And Principal Findings: SGTA consists of an N-terminal dimerisation domain (SGTA_NT), a central tetratricopeptide repeat (TPR) domain, and a glutamine rich region towards the C-terminus.

A Ctf4 trimer couples the CMG helicase to DNA polymerase α in the eukaryotic replisome.

Efficient duplication of the genome requires the concerted action of helicase and DNA polymerases at replication forks to avoid stalling of the replication machinery and consequent genomic instability. In eukaryotes, the physical coupling between helicase and DNA polymerases remains poorly understood. Here we define the molecular mechanism by which the yeast Ctf4 protein links the Cdc45-MCM-GINS (CMG) DNA helicase to DNA polymerase α (Pol α) within the replisome.

Structure of the Sgt2/Get5 complex provides insights into GET-mediated targeting of tail-anchored membrane proteins.

Small, glutamine-rich, tetratricopeptide repeat protein 2 (Sgt2) is the first known port of call for many newly synthesized tail-anchored (TA) proteins released from the ribosome and destined for the GET (Guided Entry of TA proteins) pathway. This leads them to the residential membrane of the endoplasmic reticulum via an alternative to the cotranslational, signal recognition particle-dependent mechanism that their topology denies them. In yeast, the first stage of the GET pathway involves Sgt2 passing TA proteins on to the Get4/Get5 complex through a direct interaction between the N-terminal (NT) domain of Sgt2 and the ubiquitin-like (UBL) domain of Get5.

1H, 13C and 15N assignments of Sgt2 N-terminal dimerisation domain and its binding partner, Get5 Ubiquitin-like domain.

The first stage of the GET (guided entry of tail-anchored proteins) mechanism for tail-anchored (TA) membrane protein insertion is thought to occur when Sgt2 (small, glutamine-rich, tetratricopeptide repeat-containing protein 2) binds TA proteins upon their release from the ribosome. It sorts them and passes the majority over to a complex of Get5 and Get4 for transmission along the GET pathway and delivery to their membrane destination. Sgt2 is a 38 kDa protein consisting of three domains.

Spatial organization of the mouse genome and its role in recurrent chromosomal translocations.

The extent to which the three-dimensional organization of the genome contributes to chromosomal translocations is an important question in cancer genomics. We generated a high-resolution Hi-C spatial organization map of the G1-arrested mouse pro-B cell genome and used high-throughput genome-wide translocation sequencing to map translocations from target DNA double-strand breaks (DSBs) within it. RAG endonuclease-cleaved antigen-receptor loci are dominant translocation partners for target DSBs regardless of genomic position, reflecting high-frequency DSBs at these loci and their colocalization in a fraction of cells.

Structural basis of differential ligand recognition by two classes of bis-(3'-5')-cyclic dimeric guanosine monophosphate-binding riboswitches.

The bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP) signaling pathway regulates biofilm formation, virulence, and other processes in many bacterial species and is critical for their survival. Two classes of c-di-GMP-binding riboswitches have been discovered that bind this second messenger with high affinity and regulate diverse downstream genes, underscoring the importance of RNA receptors in this pathway. We have solved the structure of a c-di-GMP-II riboswitch, which reveals that the ligand is bound as part of a triplex formed with a pseudoknot.