Biology subjects

Lloyd, R. S.

Publications and source records attributed to Lloyd, R. S..

4 recordsLinked to original sources

bioRxiv2026

Small molecule agonists of 8-oxoguanine DNA glycosylase, OGG1

Base excision repair (BER) is the primary pathway that removes oxidatively-induced DNA base damage from the nuclear and mitochondrial genomes, with 8-oxoguanine DNA glycosylase (OGG1) initiating repair at the two most frequently-formed base lesions: 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxoGua) and 2,6-diamino-4-oxo-5-formamidopyrimidine (FapyGua). Humans expressing a catalytically-compromised variant of OGG1 (S326C) are at increased risk for type 2 diabetes, Alzheimers disease, and Parkinsons disease. To potentially enhance the overall catalytic efficiency of this variant, a prior medicinal chemistry screen discovered seven chemically distinct agonists of OGG1 that stimulated activity in vitro and attenuated a paraquat (PQ) challenge in cultured cells. Herein, we developed structure-activity relationships around one specific core structure, F01. Using fluorescence-based DNA cleavage assays, we assessed the abilities of these compounds to stimulate the overall rate of OGG1 catalysis. Multiple compounds were identified that increased OGG1 activity on DNAs containing a site-specific 8-oxoGua by 2-fold or greater, with 9 compounds showing EC50 concentrations lower than F01 and were specific for OGG1. Selected agonists were shown to enhance OGG1-catalyzed release of 8-oxoGua and FapyGua from {gamma}-irradiated high-molecular-weight DNA using gas chromatography tandem mass spectrometry analyses. Since these assays did not reveal which step in the overall reaction was stimulated, we used a separation-of-function OGG1 mutant that possessed glycosylase, but not abasic-site (AP) lyase activity to demonstrate that the glycosylase step was not enhanced. In contrast, all agonists stimulated the AP lyase activity to levels equal to or greater than the magnitude of stimulation observed for overall chemistry, implicating agonist-mediated turnover as a potential contributor to the overall rate stimulation. The biological activities of selected agonists were evaluated in OGG1-deficient Kasumi-1 cells under conditions of paraquat (PQ)-induced oxidative stress, with several compounds mitigating PQ challenge.

biochemistry↗

bioRxiv2025

In vitro evidence for bisphenol A as a human liver carcinogen: Environmentally relevant doses inhibit cancer protective ESR1 signaling in a human liver cell line

I.Several environmentally ubiquitous endocrine disrupting chemicals (EDCs) are suspected carcinogens, but their mechanism(s) of action are unknown. In this study, we test the potential for a model EDC, bisphenol A (BPA), to both initiate (via oxidative mutagenesis) and promote (via endocrine disruption) liver carcinogenesis. This study is motivated by our prior finding that developmental BPA exposure caused hepatocellular carcinoma (HCC) in mice. Here, we provide in vitro evidence supporting a mechanism for BPA as a non-genotoxic carcinogen. Using a highly sensitive, error-corrected sequencing method, we demonstrate that human population-relevant doses of BPA cause mutations that are consistent with oxidative DNA damage; however, overall mutation frequency does not differ substantially from controls. In contrast, we show that BPA inhibits cancer-protective, estrogen-induced transcription of estrogen receptor 1 (ESR1) target genes in the presence of pre-pubertal, but not post-pubertal, levels of estradiol. These results constitute strong initial evidence supporting BPA as a liver cancer promoting agent. This mechanism may be generalizable to a wide range of environmental EDCs that are weak agonists for ESR1. This finding is critically important to prevention of HCC, which is prevalent, lethal, and poorly responsive to therapy.

pharmacology and toxicology↗

bioRxiv2023

Spontaneous allelic variant in Ush1g resulting in an expanded phenotype

Strategies to reveal the discovery of the relationships between novel phenotypic behaviors and specific genetic alterations can be achieved via either target-specific, directed mutagenesis or phenotypic selection following random chemical mutagenesis. As an alternative approach, one can exploit deficiencies in DNA repair pathways that are responsible for the maintenance of genetic integrity in response to spontaneously-induced damage. In the genetic background of mice deficient in the DNA glycosylase NEIL1, elevated numbers of spontaneous mutations arise from translesion DNA synthesis past unrepaired, oxidatively-induced base damage. Several litters of Neil1 knockout mice included animals that were distinguished by their backwards-walking behavior in open-field environments, while maintaining frantic forward movements in their home cage environment. Other phenotypic manifestations included swim test failures, head tilting, and circling. Mapping of the mutation that conferred these behaviors revealed the introduction of a stop codon at amino acid 4 of the Ush1g gene; the allele was Ush1gbw, reflecting the backwards-walking phenotype. Ush1gbw/bw null mice displayed auditory and vestibular defects that are commonly seen with mutations affecting inner-ear hair-cell function, including a complete lack of auditory brainstem responses and vestibular-evoked potentials. As in other Usher syndrome type I mutant mouse lines, hair-cell phenotypes included disorganized and split hair bundles, as well as altered distribution of proteins for stereocilia that localize to the tips of row 1 or row 2. Disruption to the bundle and kinocilium displacement suggested that USH1G is essential for forming the hair cells kinocilial links. Due to the vestibular dysfunction, however, visual behavior as measured with optokinetic tracking could not be assessed in Ush1gbw/bw mice. Consistent with other Usher type 1 models, however, Ush1gbw/bw mice had no substantial retinal degeneration compared to Ush1gbw/+ controls out to six months. In contrast to previously-described Ush1g alleles, this new allele provides the first knockout model for this gene.

neuroscience↗

bioRxiv2017

Ectopic Expression of DNA Repair Enzymes Modulates Survival following Ultraviolet Irradiation Challenge

In Escherichia coli, the nucleotide excision repair (NER) pathway removes ultraviolet (UV) light-induced cyclobutane pyrimidine dimers (CPDs) and 6-4 dipyrimidine photoproducts (6-4 PPs). Activation of alternative repair pathways, such as base excision repair (BER) and nucleotide incision repair (NIR), is inoperative because this organism lacks both the necessary BER DNA glycosylase and NIR UV endonuclease to initiate repair of these lesions. To determine if initiation of either pathway would enhance survival to biologically-relevant UV irradiation, the BER and NIR pathways were activated by expression of Chlorella virus-1 pyrimidine dimer glycosylase (cv-pdg) and Schizosaccharomyces pombe UV endonuclease (UVDE), respectively. The substrate specificity of cv-pdg includes CPDs and ring-fragmented purines, 4,6-diamino-5-formamidopyrimidine and 2,6-diamino-4-hydroxy-5-formamidopyrimidine, but not 6-4 PPs. In contrast, while UVDE incises DNA containing CPDs and 6-4 PPs, it was not previously known if the substrate specificity of UVDE included DNA containing ring-fragmented purines. Mass spectrometry was used to establish that these oxidatively-induced lesions were not substrates for UVDE. Expression of either cv-pdg or UVDE in NER-deficient E. coli significantly enhanced survival following UVB irradiation, but not to the levels of wild type (WT) cells. Survival of NER-proficient, homologous recombination-deficient cells could also be significantly enhanced by expression of either enzyme, suggesting that in response to UVB exposure, interactions between NER and activated BER or NIR pathways could be additive. Further, expression of cv-pdg or UVDE in WT E. coli enhanced survival following solar-simulated light (SSL) exposures.

microbiology↗