RESUMO
DNA polymerase stalling activates the ATR checkpoint kinase, which in turn suppresses fork collapse and breakage. Herein, we describe use of ATR inhibition (ATRi) as a means to identify genomic sites of problematic DNA replication in murine and human cells. Over 500 high-resolution ATR-dependent sites were ascertained using two distinct methods: replication protein A (RPA)-chromatin immunoprecipitation (ChIP) and breaks identified by TdT labeling (BrITL). The genomic feature most strongly associated with ATR dependence was repetitive DNA that exhibited high structure-forming potential. Repeats most reliant on ATR for stability included structure-forming microsatellites, inverted retroelement repeats, and quasi-palindromic AT-rich repeats. Notably, these distinct categories of repeats differed in the structures they formed and their ability to stimulate RPA accumulation and breakage, implying that the causes and character of replication fork collapse under ATR inhibition can vary in a DNA-structure-specific manner. Collectively, these studies identify key sources of endogenous replication stress that rely on ATR for stability.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/genética , Replicação do DNA/genética , Repetições de Microssatélites/genética , Animais , Proteínas de Ciclo Celular/genética , Cromatina/genética , Imunoprecipitação da Cromatina/métodos , Quebras de DNA de Cadeia Dupla , Dano ao DNA/genética , Feminino , Instabilidade Genômica/genética , Humanos , Camundongos , Proteína de Replicação A/genéticaRESUMO
Here we present the case of a patient who purchased a Hawthorne root (Crataegus mexicana) product, Raiz de Tejocote, for weight loss purposes. She presented with diffuse myalgias, dizziness and a heart rate of 52 beats per minute. At triage and at initial evaluation, the patient denied taking any medications, but on iterative questioning concerning over-the-counter, over-the-internet and herbal medications, she reported taking Hawthorne root tablets in the three days prior to the emergency department (ED) visit for the purpose of weight loss. The product was purchashed through the internet. Her plasma digoxin concentration was 0.4 ng/ml the patient's constellation of symptoms, as well as the detectable plasma digoxin concentration, were consistent with hawthorne root toxicity. Hawthorne root has intrinsic cardiac glycoside activity. In addition, Hawthorne root may cause a range of toxicity. Mild symptoms can include flu-like syndrome with significant myalgias. However, in the more severe exposures the cardiac glycoside effects can result in bradycardia and hemodynamic instability. Symptoms resolved with ED observation. The heart rate normalized. This case reinforces the importance of asking a patient about all medications, including over-the-counter, over-the-internet and herbal medications.