ssDNA reeling is an intermediate step in the reiterative DNA unwinding activity of the WRN-1 helicase.

RecQ helicases are highly conserved between bacteria and humans. These helicases unwind various DNA structures in the 3' to 5'. Defective helicase activity elevates genomic instability and is associated with predisposition to cancer and/or premature aging.

Molecular characteristics of reiterative DNA unwinding by the Caenorhabditis elegans RecQ helicase.

The RecQ family of helicases is highly conserved both structurally and functionally from bacteria to humans. Defects in human RecQ helicases are associated with genetic diseases that are characterized by cancer predisposition and/or premature aging. RecQ proteins exhibit 3'-5' helicase activity and play critical roles in genome maintenance.

Multiple RPAs make WRN syndrome protein a superhelicase.

RPA is known to stimulate the helicase activity of Werner syndrome protein (WRN), but the exact stimulation mechanism is not understood. We use single-molecule FRET and magnetic tweezers to investigate the helicase activity of WRN and its stimulation by RPA. We show that WRN alone is a weak helicase which repetitively unwind just a few tens of base pairs, but that binding of multiple RPAs to the enzyme converts WRN into a superhelicase that unidirectionally unwinds double-stranded DNA more than 1 kb.

Active Control of Repetitive Structural Transitions between Replication Forks and Holliday Junctions by Werner Syndrome Helicase.

The reactivation of stalled DNA replication via fork regression invokes Holliday junction formation, branch migration, and the recovery of the replication fork after DNA repair or error-free DNA synthesis. The coordination mechanism for these DNA structural transitions by molecular motors, however, remains unclear. Here we perform single-molecule fluorescence experiments with Werner syndrome protein (WRN) and model replication forks.

DNA Strand Breaks in Mitotic Germ Cells of Caenorhabditis elegans Evaluated by Comet Assay.

DNA damage responses are important for the maintenance of genome stability and the survival of organisms. Such responses are activated in the presence of DNA damage and lead to cell cycle arrest, apoptosis, and DNA repair. In Caenorhabditis elegans, double-strand breaks induced by DNA damaging agents have been detected indirectly by antibodies against DSB recognizing proteins.

The Caenorhabditis elegans Werner syndrome protein participates in DNA damage checkpoint and DNA repair in response to CPT-induced double-strand breaks.

The RecQ helicases play roles in maintenance of genomic stability in species ranging from Escherichia coli to humans and interact with proteins involved in DNA metabolic pathways such as DNA repair, recombination, and replication. Our previous studies found that the Caenorhabditis elegans WRN-1 RecQ protein (a human WRN ortholog) exhibits ATP-dependent 3'-5' helicase activity and that the WRN-1 helicase is stimulated by RPA-1 on a long forked DNA duplex. However, the role of WRN-1 in response to S-phase associated with DSBs is unclear.

WRN translocation from nucleolus to nucleoplasm is regulated by SIRT1 and required for DNA repair and the development of chemoresistance.

When defective or absent, Werner syndrome protein (WRN) causes a genetic premature aging disorder called Werner syndrome. Several studies have reported that defects in WRN function are responsible for not only progeria syndrome but also genomic instability via the deregulation of DNA repair, replication, recombination, and telomere stability. Given the importance of WRN in the repair process, we herein investigated the potential role of WRN in drug response by evaluating the DNA repair following exposure to cisplatin in human cancer cell lines.

Characterization of the Caenorhabditis elegans HIM-6/BLM helicase: unwinding recombination intermediates.

Mutations in three human RecQ genes are implicated in heritable human syndromes. Mutations in BLM, a RecQ gene, cause Bloom syndrome (BS), which is characterized by short stature, cancer predisposition, and sensitivity to sunlight. BLM is a RecQ DNA helicase that, with interacting proteins, is able to dissolve various DNA structures including double Holliday junctions.

Cockayne Syndrome group B protein stimulates NEIL2 DNA glycosylase activity.

Cockayne Syndrome is a segmental premature aging syndrome, which can be caused by loss of function of the CSB protein. CSB is essential for genome maintenance and has numerous interaction partners with established roles in different DNA repair pathways including transcription coupled nucleotide excision repair and base excision repair. Here, we describe a new interaction partner for CSB, the DNA glycosylase NEIL2.

Physical and functional interactions of Caenorhabditis elegans WRN-1 helicase with RPA-1.

The Caenorhabditis elegans Werner syndrome protein, WRN-1, a member of the RecQ helicase family, has a 3'-5' DNA helicase activity. Worms with defective wrn-1 exhibit premature aging phenotypes and an increased level of genome instability. In response to DNA damage, WRN-1 participates in the initial stages of checkpoint activation in concert with C.

DNA binding and unwinding by self-assembled supramolecular hetero-bimetallacycles.

Two new tetracationic hetero-bimetallacycles were prepared from a bis-pyridine amide ligand and metal (Pd and Pt) acceptors. We found that both self-assembled hetero-bimetallacycles bind and unwind supercoiled DNA as established by photophysical and gel electrophoresis analyses, respectively.

DNA secondary structure of the released strand stimulates WRN helicase action on forked duplexes without coordinate action of WRN exonuclease.

Werner syndrome (WS) is an autosomal recessive premature aging disorder characterized by aging-related phenotypes and genomic instability. WS is caused by mutations in a gene encoding a nuclear protein, Werner syndrome protein (WRN), a member of the RecQ helicase family, that interestingly possesses both helicase and exonuclease activities. Previous studies have shown that the two activities act in concert on a single substrate.

The Caenorhabditis elegans Werner syndrome protein functions upstream of ATR and ATM in response to DNA replication inhibition and double-strand DNA breaks.

WRN-1 is the Caenorhabditis elegans homolog of the human Werner syndrome protein, a RecQ helicase, mutations of which are associated with premature aging and increased genome instability. Relatively little is known as to how WRN-1 functions in DNA repair and DNA damage signaling. Here, we take advantage of the genetic and cytological approaches in C.

The structure of the trimer of human 4-1BB ligand is unique among members of the tumor necrosis factor superfamily.

Binding of the 4-1BB ligand (4-1BBL) to its receptor, 4-1BB, provides the T lymphocyte with co-stimulatory signals for survival, proliferation, and differentiation. Importantly, the 4-1BB-4-1BBL pathway is a well known target for anti-cancer immunotherapy. Here we present the 2.

Mechanism of Werner DNA helicase: POT1 and RPA stimulates WRN to unwind beyond gaps in the translocating strand.

WRN belongs to the RecQ family of DNA helicases and it plays a role in recombination, replication, telomere maintenance and long-patch base excision repair. Here, we demonstrate that WRN efficiently unwinds DNA substrates containing a 1-nucleotide gap in the translocating DNA strand, but when the gap size is increased to 3-nucleotides unwinding activity significantly declines. In contrast, E.

Roles of RECQ helicases in recombination based DNA repair, genomic stability and aging.

The maintenance of the stability of genetic material is an essential feature of every living organism. Organisms across all kingdoms have evolved diverse and highly efficient repair mechanisms to protect the genome from deleterious consequences of various genotoxic factors that might tend to destabilize the integrity of the genome in each generation. One such group of proteins that is actively involved in genome surveillance is the RecQ helicase family.

Genotype-phenotype relationship between DNA repair gene genetic polymorphisms and DNA repair capacity.

Genotype-phenotype relationships between genetic polymorphisms of DNA repair genes and DNA repair capacity were evaluated in a case-control study of breast cancer. Selected DNA repair genes included were those involved in double-strand break repair (ATM, XRCC2, XRCC4, XRCC6, LIG4, RAD51, RAD52), base excision repair (LIG1), nucleotide excision repair (ERCC1), and mismatch repair (hMLH1). The subjects consisted of histologically confirmed breast cancer cases (n=132) and controls (n=75) with no present or previous history of cancer.

Biochemical characterization of the WRN-1 RecQ helicase of Caenorhabditis elegans.

The highly conserved RecQ helicases are essential for the maintenance of genomic stability. Werner syndrome protein, WRN, is one of five human RecQ helicase homologues, and a deficiency of the protein causes a hereditary premature aging disorder that is characterized by genomic instability. A WRN orthologue, wrn-1 lacking the exonuclease domain, has been identified in the nematode Caenorhabditis elegans.

Longevity and resistance to stress correlate with DNA repair capacity in Caenorhabditis elegans.

DNA repair is an important mechanism by which cells maintain genomic integrity. Decline in DNA repair capacity or defects in repair factors are thought to contribute to premature aging in mammals. The nematode Caenorhabditis elegans is a good model for studying longevity and DNA repair because of key advances in understanding the genetics of aging in this organism.

Werner syndrome protein participates in a complex with RAD51, RAD54, RAD54B and ATR in response to ICL-induced replication arrest.

Werner syndrome (WS) is a rare genetic disorder characterized by genomic instability caused by defects in the WRN gene encoding a member of the human RecQ helicase family. RecQ helicases are involved in several DNA metabolic pathways including homologous recombination (HR) processes during repair of stalled replication forks. Following introduction of interstrand DNA crosslinks (ICL), WRN relocated from nucleoli to arrested replication forks in the nucleoplasm where it interacted with the HR protein RAD52.

Crystallization and preliminary X-ray crystallographic study of the extracellular domain of the 4-1BB ligand, a member of the TNF family.

The 4-1BB ligand, a member of the tumour necrosis factor (TNF) family, is an important co-stimulatory molecule that plays a key role in the clonal expansion and survival of CD8+ T cells. Signalling through binding of the 4-1BB ligand and 4-1BB has been reported to enhance CD8+ T-cell expansion and protect activated CD8+ T cells from death. The 4-1BB ligand is an integral protein expressed on activated antigen-presenting cells.

Genetic polymorphisms of ataxia telangiectasia mutated and breast cancer risk.

To evaluate the role of genetic polymorphisms of ataxia telangiectasia mutated (ATM) in the etiology of breast cancer, a hospital-based case-control study was conducted in Korea. Nine-hundred ninety-six histologically confirmed incident breast cancer cases and 1,181 cancer-free controls were recruited in Seoul between 1995 and 2003. Genotypes of the ATM polymorphisms-5144A > T, IVS21 + 1049T > C, IVS33 - 55T > C, IVS34 + 60G > A, and 3393T > G were determined by the 5'-nuclease assay.

Repair of mitomycin C cross-linked DNA in mammalian cells measured by a host cell reactivation assay.

DNA repair capacity in a cell could be detected by a host-cell reactivation assay (HCR). Since relation between DNA repair and genetic susceptibility to cancer remains unclear, it is necessary to identify DNA repair defects in human cancer cells. To assess DNA repair for breast cancer susceptibility, we developed a modified HCR assay using a plasmid containing a firefly luciferase gene damaged by mitomycin C (MMC), which forms interstrand cross-link (ICL) adducts.

Regulation of WRN helicase activity in human base excision repair.

Werner syndrome patients are deficient in the Werner protein (WRN), which is a multifunctional nuclear protein possessing 3'-5' exonuclease and ATP-dependent helicase activities. Studies of Werner syndrome cells and biochemical studies of WRN suggest that WRN plays a role in several DNA metabolic pathways. WRN interacts with DNA polymerase beta (pol beta) and stimulates pol beta strand displacement synthesis on a base excision repair (BER) intermediate in a helicase-dependent manner.

Chromatin remodeling facilitates DNA incision in UV-damaged nucleosomes.

The DNA repair machinery must locate and repair DNA damage all over the genome. As nucleosomes inhibit DNA repair in vitro, it has been suggested that chromatin remodeling might be required for efficient repair in vivo. To investigate a possible contribution of nucleosome dynamics and chromatin remodeling to the repair of UV-photoproducts in nucleosomes, we examined the effect of a chromatin remodeling complex on the repair of UV-lesions by Micrococcus luteus UV endonuclease (ML-UV endo) and T4-endonuclease V (T4-endoV) in reconstituted mononucleosomes positioned at one end of a 175-bp long DNA fragment.