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Androgen receptor (AR) signalling is essential in nearly all prostate cancers. Any alterations to AR-mediated transcription can have a profound effect on carcinogenesis and tumor growth. While mutations of the AR protein have been extensively studied, little is known about those somatic mutations that occur at the non-coding regions where AR binds DNA. Using clinical whole genome sequencing, we show that AR binding sites have a dramatically increased rate of mutations that is greater than any other transcription factor and specific to only prostate cancer. Demonstrating this may be common to lineage-specific transcription factors, estrogen receptor binding sites were also found to have elevated rate of mutations in breast cancer. We provide evidence that these mutations at AR binding sites, and likely other related transcription factors, are caused by faulty repair of abasic sites. Overall, this work demonstrates that non-coding AR binding sites are frequently mutated in prostate cancer and can impact enhancer activity. Androgen receptor (AR) mediated transcription is critical to prostate tumorigenesis and development. Here, utilising clinical whole genome sequencing data, the authors show that the non-coding AR binding sites on DNA are frequently mutated in prostate cancer potentially due to faulty base excision repair mechanisms

APE1 is the major nuclease for excising abasic (AP) sites and particular 3'-obstructive termini from DNA, and is an integral participant in the base excision repair (BER) pathway. BER capacity plays a prominent role in dictating responsiveness to agents that generate oxidative or alkylation DNA damage, as well as certain chain-terminating nucleoside analogs and 5-fluorouracil. We describe within the development of a robust, 1536-well automated screening assay that employs a deoxyoligonucleotide substrate operating in the red-shifted fluorescence spectral region to identify APE1 endonuclease inhibitors. This AP site incision assay was used in a titration-based high-throughput screen of the Library of Pharmacologically Active Compounds (LOPAC(1280)), a collection of well-characterized, drug-like molecules representing all major target classes. Prioritized hits were authenticated and characterized via two high-throughput screening assays -- a Thiazole Orange fluorophore-DNA displacement test and an E. coli endonuclease IV counterscreen -- and a conventional, gel-based radiotracer incision assay. The top, validated compounds, i.e. 6-hydroxy-DL-DOPA, Reactive Blue 2 and myricetin, were shown to inhibit AP site cleavage activity of whole cell protein extracts from HEK 293T and HeLa cell lines, and to enhance the cytotoxic and genotoxic potency of the alkylating agent methylmethane sulfonate. The studies herein report on the identification of novel, small molecule APE1-targeted bioactive inhibitor probes, which represent initial chemotypes towards the development of potential pharmaceuticals.

Base excision repair (BER) is the predominant pathway for coping with most forms of hydrolytic, oxidative or alkylative DNA damage. Measuring BER capacity in living cells is valuable for both basic science applications and epidemiological studies, since deficiencies in this pathway have been associated with cancer susceptibility and other adverse health outcomes. At present, there is an ongoing effort to develop methods to effectively quantify the rate of BER as a whole. We present a variation of a previously described “Oligonucleotide Retrieval Assay” designed to measure DNA excision repair that is capable of quantifying the rate of repair of thymine glycol in a variety of human cells with a high degree of sensitivity.

Cockayne syndrome (CS) is an inherited disorder that involves photosensitivity, developmental defects, progressive degeneration and characteristics of premature aging. Evidence indicates primarily nuclear roles for the major CS proteins, CSA and CSB, specifically in DNA repair and RNA transcription. We reveal herein a complex regulation of CSB targeting that involves three major consensus signals: NLS1 (aa467-481), which directs nuclear and nucleolar localization in cooperation with NoLS1 (aa302-341), and NLS2 (aa1038-1055), which seemingly optimizes nuclear enrichment. CSB localization to the nucleolus was also found to be important for full UVC resistance. CSA, which does not contain any obvious targeting sequences, was adversely affected (i.e. presumably destabilized) by any form of truncation. No inter-coordination between the subnuclear localization of CSA and CSB was observed, implying that this aspect does not underlie the clinical features of CS. The E3 ubiquitin ligase binding partner of CSA, DDB1, played an important role in CSA stability (as well as DDB2), and facilitated CSA association with chromatin following UV irradiation; yet did not affect CSB chromatin binding. We also observed that initial recruitment of CSB to DNA interstrand crosslinks is similar in the nucleoplasm and nucleolus, although final accumulation is greater in the former. Whereas assembly of CSB at sites of DNA damage in the nucleolus was not affected by RNA polymerase I inhibition, stable retention at these sites of presumed repair was abrogated. Our studies reveal a multi-faceted regulation of the intranuclear dynamics of CSA and CSB that plays a role in mediating their cellular functions.

Background: To adapt and validate cross-culturally the Oral Health Values (OHVS) questionnaire to Portuguese language. Methods: The OHVS questionnaire was culturally translated and adapted according to international guidelines. We enrolled 280 patients in a population-based epidemiological survey conducted at the Egas Moniz Dental Clinic (Almada, Portugal). The participants answered the Portuguese version of the OHVS (OHVS-PT), which is a 12-item scale with four-factor structure (Professional Dental Care, Appearance and Health, Flossing and Retaining Natural Teeth factor). Psychometric properties were tested using content validity, construct validity, internal consistency, and test–retest reliability. Results: The OHVS-PT presented adequate reliability (ICC = 0.93, 95% confidence interval (CI): 0.86; 0.97, p < 0.001) with values for the Cronbach’s alpha coefficient of the sub-constructs ranging from 0.92 to 0.98. In the Confirmatory Factor Analysis, the final models presented good fit, with the Comparative Fit Indices ranging from 0.882 to 0.891 and the root-mean-squared error of Approximation between 0.065 and 0.069. Conclusions: The OHVS-PT was shown to be a valid and reliable tool to assess oral health values in a Portuguese population. Further studies should evaluate the psychometric properties of the oral personal representation on dental specialties and its impact on dental appointments and procedures.

Many environmental metals are co-carcinogens, eliciting their effects via inhibition of DNA repair. Apurinic/apyrimidinic (AP) endonuclease 1 (Ape1) is the major mammalian abasic endonuclease and initiates repair of this cytotoxic/mutagenic lesion by incising the DNA backbone via a Mg2+-dependent reaction. In this study we examined the effects of arsenite [As(III)], cadmium [Cd(II)], cobalt [Co(II)], iron [Fe(II)], nickel [Ni(II)], and lead [Pb(II)] at concentrations ranging from 0.3 to 100 μM on the incision activity of Apel in the presence of 1 mM MgCl2. Pb(II) and Fe(II) inhibited Apel activity at each of the concentrations tested, with an IC50(half-maximal inhibitory concentration) of 0.61 and 1.0 μM, respectively. Cd(II) also inhibited Apel activity but only at concentrations > 10 μM. No inhibition was seen with As(III), Co(II), or Ni(II). A similar inhibition pattern was observed with the homologous Escherichia coli protein, exonuclease III, but no inhibition was seen with the structurally distinct AP endonuclease E. coli endonuclease IV, indicating a targeted effect of Pb(II), Fe(II), and Cd(II) on the Apel-like repair enzymes. Excess nonspecific DNA did not abrogate the metal inactivation, suggesting a protein-specific effect. Notably, Cd(II), Fe(II), and Pb(II) [but not As(III), Co(II), or Ni(II)] inhibited AP endonuclease activity in whole-cell extracts but had no significant effect on single nucleotide gap filling, 5′-flap endonuclease, and nick ligation activities, supporting the idea of selective inactivation of Apel in cells. Our results are the first to identify a potential DNA repair enzyme target for lead and suggest a means by which these prevalent environmental metals may elicit their deleterious effects.

Throughout history, Indigenous people have stewarded seascapes, with people–nature reciprocity being a key component of many Indigenous management strategies, yet it is often overlooked in fisheries management. This research focuses on northern abalone (Haliotis kamtschatkana) in Haida Gwaii, the home of the Haida Nation. The Haida have developed a range of values associated with abalone, including reciprocity. However, commercial fishing and colonial mismanagement have led to overexploitation, resulting in a prohibition on all gatherings since 1990. Despite this, the relationship between the Haida and abalone persists in the community's knowledge. We used the Intergovernmental Science‐Policy Platform on Biodiversity and Ecosystem Services (IPBES) framework and the concept of reciprocal contributions to help paint a complex relationship between Haida knowledge holders and abalone, identifying values, benefits, drivers and barriers. We identified food uses and meanings that include emotional experiences. Additionally, we compiled an array of reciprocal contributions, such as enhancement actions, fishing guidelines and nature protection practices. These memories and guidelines have been passed down through generations. The principle of ‘take only what you need to eat’ is essential for managing abalone populations. Reciprocal actions have been crucial for both abalone and the Haida. Elders have transplanted abalone to other sites, and the Council of the Haida Nation continues monitoring and restoration work. Cultural and ecological education about abalone is preserved through family stories, school activities and ceremonies, reinforcing its status as a culturally important species. The abalone fishing boom and Canadian mismanagement were identified as negative drivers that significantly impacted abalone populations. Most Haida elders recall their efforts in the 1970s to raise concerns with the federal fishing agency about the declining abalone population, but their warnings went unheeded. Currently, due to limited enforcement capacity, abalone poaching is the primary concern, while bureaucratic hurdles in decision‐making processes impede effective assessment and management. This work contributes to reviewing the past and rethinking management strategies in local contexts where values like reciprocity remain integral to Indigenous people. It also aims to generate dialogues and actions with international fisheries platforms, facilitating learning from other local experiences and recognizing common colonial barriers worldwide. Read the free Plain Language Summary for this article on the Journal blog. Read the free Plain Language Summary for this article on the Journal blog.

Nitrosative Stress, Uric Acid, and Peripheral Nerve Function in Early Type 1 Diabetes Robert D. Hoeldtke 1 , Kimberly D. Bryner 1 , Daniel R. McNeill 1 , Gerald R. Hobbs 2 , Jack E. Riggs 3 , Sarah S. Warehime 1 , Ian Christie 4 , Gary Ganser 4 and Knox Van Dyke 5 1 Department of Medicine, West Virginia University (WVU), Morgantown, West Virginia 2 Department of Community Medicine and Statistics, WVU, Morgantown, West Virginia 3 Department of Neurology, WVU, Morgantown, West Virginia 4 Department of Mathematics, WVU, Morgantown, West Virginia 5 Department of Biochemistry and Molecular Pharmacology, WVU, Morgantown, West Virginia Abstract The present study was performed to determine whether nitric oxide overproduction is associated with deterioration in peripheral nerve function in type 1 diabetes. We measured peripheral nerve function and biochemical indicators of nitrosative stress annually for 3 years in 37 patients with type 1 diabetes. Plasma nitrite and nitrate (collectively NO x ) were 34.0 ± 4.9 μmol/l in the control subjects and 52.4 ± 5.1, 50.0 ± 5.1, and 49.0 ± 5.2 in the diabetic patients at the first, second, and third evaluations, respectively ( P < 0.01). Nitrotyrosine (NTY) was 13.3 ± 2.0 μmol/l in the control subjects and 26.8 ± 4.4, 26.1 ± 4.3, and 32.7 ± 4.3 in the diabetic patients ( P < 0.01). Uric acid was suppressed by 20% in the diabetic patients ( P < 0.001). Composite motor nerve conduction velocity for the median, ulnar, and peroneal nerves was decreased in patients with high versus low NTY (mean Z score −0.522 ± 0.25 versus 0.273 ± 0.22; P < 0.025). Patients with high NO x had decreased sweating, and those with suppressed uric acid had decreased autonomic function. In conclusion, nitrosative stress in early diabetes is associated with suppressed uric acid and deterioration in peripheral nerve function. Footnotes Address correspondence and reprint requests to Robert D. Hoeldtke, West Virginia University School of Medicine, Department of Medicine, One Medical Center Drive, PO Box 9159, Morgantown, WV 26506-9159. Received for publication 3 April 2002 and accepted in revised form 28 May 2002. 8-iso-PGF2α, 8-isoprostaglandin F2α; ELISA, enzyme-linked immunosorbent assay; eNOS, endothelial nitric oxide synthase; HPLC, high-performance liquid chromatography; iNOS, inducible nitric oxide synthase; NTY, nitrotyrosine; TY, tyrosine; VMA, vanillylmandelic acid. DIABETES