OverviewWerner syndrome protein (WRN) is one of a family of five human RecQ helicases implicated in the maintenance of genome stability. The conserved RecQ family also includes RecQ1, Bloom syndrome protein (BLM), RecQ4, and RecQ5 in humans (see additional reviews in this issue), as well as Sgs1 in Saccharomyces cerevisiae, Rqh1 in Schizosaccharomyces pombe, and homologs in Caenorhabditis elegans, Xenopus laevis, and Drosophila melanogaster [1]. Defects in three of the RecQ helicases, RecQ4, BLM, and WRN, cause human pathologies linked with cancer predisposition and premature aging [1][2][3][4][5]. Mutations in the WRN gene are the causative factor of Werner syndrome (WS). WRN is one of the best characterized of the RecQ helicases and is known to have roles in DNA replication and repair, transcription, and telomere maintenance [1][2][3][4][5][6]. Studies both in vitro and in vivo indicate that the roles of WRN in a variety of DNA processes are mediated by post-translational modifications, as well as several important protein-protein interactions [1,2,7]. Many of these functions of WRN in genome maintenance, as well as the clinical characteristics of WS, have been recently reviewed [8][9][10][11][12]. In this work, we will summarize some of the early studies on the cellular roles of WRN and highlight the recent findings that shed some light on the link between the protein and its cellular functions with the disease pathology. Molecular Genetics of Werner Syndrome (WS)WS is a rare autosomal recessive progeroid disorder characterized by the development of cataracts, changing of skin conditions, bird-like facies, atypical short stature, and premature graying or thinning of the hair [9]. Patients also often develop hypogonadism, osteoporosis, diabetes mellitus, artherosclerosis [13], and cancers, particularly sarcomas [14]. Onset of symptoms usually occurs in the third decade of life, and health subsequently declines with median age at death between 47-54 years [13]. Because WS presents with early-onset of conditions commonly seen in the aged, it is a good model system for the study of mechanisms of normal aging [15,16]. WRN BiochemistryThe causative factor of the majority of WS is mutation in the WRN gene, which codes for a member of the highly conserved RecQ family of helicases. While several different mutations within the gene are seen in WS, most result in production of truncated WRN protein [13]. 1Address correspondence to: Dr. Vilhelm A. Bohr, NIH Biomedical Research Center, 251 Bayview Boulevard, Baltimore, MD 21224; BohrV@grc.nia.nih.gov. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the...

Background: RECQL4 is a RecQ helicase mutated in Rothmund-Thomson Syndrome (RTS) and has a functional role in DNA replication and repair. Results: RECQL4-depleted and RTS patient cells show telomere abnormalities and that RECQL4 interacts with telomeric DNA and related proteins. Conclusion: RECQL4 is involved in telomere maintenance. Significance: The RecQ helicase RECQL4 is involved in telomere replication and maintenance. This establishes a connection between telomere function and a disease with premature aging phenotype.

Humans have five members of the well conserved RecQ helicase family: RecQ1, Bloom syndrome protein (BLM), Werner syndrome protein (WRN), RecQ4, and RecQ5, which are all known for their roles in maintaining genome stability. BLM, WRN, and RecQ4 are associated with premature aging and cancer predisposition. Of the three, RecQ4's biological and cellular roles have been least thoroughly characterized. Here we tested the helicase activity of purified human RecQ4 on various substrates. Consistent with recent results, we detected ATP-dependent RecQ4 unwinding of forked duplexes. However, our results provide the first evidence that human RecQ4's unwinding is independent of strand annealing, and that it does not require the presence of excess ssDNA. Moreover, we demonstrate that a point mutation of the conserved lysine in the Walker A motif abolished helicase activity, implying that not the N-terminal portion, but the helicase domain is solely responsible for the enzyme's unwinding activity. In addition, we demonstrate a novel stimulation of RecQ4's helicase activity by replication protein A, similar to that of RecQ1, BLM, WRN, and RecQ5. Together, these data indicate that specific biochemical activities and protein partners of RecQ4 are conserved with those of the other RecQ helicases.

The one-electron oxidation of duplex DNA generates a nucleobase radical cation (electron "hole") that migrates long distances by a hopping mechanism. The radical cation reacts irreversibly with H2O or O2 to form oxidation products (damaged bases). In normal DNA (containing the four common DNA bases), reaction occurs most frequently at guanine. However, in DNA duplexes that do not contain guanine (i.e., those comprised exclusively of A/T base pairs), we discovered that reaction occurs primarily at thymine and gives products resulting from oxidation of the T-C5 methyl group and from addition to its C5-C6 double bond. This surprising result shows that it is the relative reactivity, not the stability, of a nucleobase radical cation that determines the nature of the products formed from oxidation of DNA. A mechanism for reaction is proposed whereby a thymine radical cation may either lose a proton from its methyl group or H2O/O2 may add across its double bond. In the latter case, addition may initiate a tandem reaction that converts both thymines of a TT step to oxidation products.

Telomeres are nucleoprotein complexes at the ends of linear chromosomes in eukaryotes, and are essential in preventing chromosome termini from being recognized as broken DNA ends. Telomere shortening has been linked to cellular senescence and human aging, with oxidative stress as a major contributing factor. 7, 8-dihydro-8-oxogaunine (8-oxodG) is one of the most abundant oxidative guanine lesions, and 8-oxoguanine DNA Glycosylase (OGG1) is involved in its removal. In this study, we examined if telomeric DNA is particularly susceptible to oxidative base damage and if telomere-specific factors affect the incision of oxidized guanines by OGG1. We demonstrated that telomeric TTAGGG repeats were more prone to oxidative base damage and repaired less efficiently than non-telomeric TG repeats in vivo. We also showed that the 8-oxodGincision activity of OGG1 is similar in telomeric and non-telomeric double-stranded substrates. In addition, telomere repeat binding factors TRF1 and TRF2 do not impair OGG1 incision activity. Yet, 8-oxodG in some telomere structures (e.g., fork-opening, 3'-overhang, and D-loop) were less effectively excised by OGG1, depending upon its position in these substrates. Collectively, our data indicate that the sequence context of telomere repeats and certain telomere configurations may contribute to telomere vulnerability to oxidative DNA damage processing.