Expanding gene sequencing tool use in environmental carcinogen studies

A technique called whole exome sequencing (WES) has been successfully applied to rats to study how a fungal toxin may induce liver cancer, according to NIEHS researchers and their collaborators.

Aflatoxin B1 (AFB1) is a widely studied, potent carcinogen associated with liver cancer. Humans are exposed to aflatoxin by consuming peanuts, maize, and cereal grains contaminated with mold. Aflatoxins elicit gene mutations through the binding of their metabolites to DNA.

The researchers set out to validate the feasibility of applying WES in rats as a mammalian model for examining genetic changes associated with tumor formation triggered by chemicals. They applied WES to DNA extracted from rat liver tissue following 14- and 90-day exposure to AFB1. The carcinogen induced mutations capable of altering multiple pathways involved in the cell cycle, replication, repair, and death, thereby contributing to the development of malignant cancer. In particular, the results revealed that mutations in Mcm8, Bdp1, and Cct6a may play an important role in driving cancer formation.

According to the authors, the study demonstrates the validity of rat WES as an important molecular tool for use in investigative, translational studies that aim to shed light on the interplay among genetics, environmental influences, and disease biology.

Citation: Foley JF, Elgart B, Phadke D, Mav D, Tripodi I, Clausen N, Weick M, Gladwell W, Gerrish K, Shah R, Merrick BA. 2023. Whole exome and transcript profiling of liver following aflatoxin B1 exposure in rats. J Appl Toxicol; doi: 10.1002/jat.4463. [Online 12 Mar 2023]. (JW)

How redox stress generates mutations

The metabolic and mutational signatures of different types of oxidizing agents vary widely, according to NIEHS researchers and their collaborators.

Redox stress is triggered by an imbalance of oxidants and antioxidants in cells, and is a hallmark of cancer aging, as well as neurological and inflammatory diseases. Redox agents can directly damage DNA and cause mutations, leading to genome instability. To understand how redox stress-induced mutagenesis contributes to disease and aging, it is necessary to investigate the genetic and metabolic effects of different types of redox agents.

Toward this goal, the researchers unraveled the mutational spectra of potassium bromate in yeast single-strand DNA and compared them to mutational footprints generated by hydrogen peroxide and paraquat. The three redox agents — potassium bromate, hydrogen peroxide, and paraquat — are used interchangeably in research, including studies in human cells. Using a combination of genetic, biochemical, biophysical, and computational approaches, the researchers unexpectedly discovered distinct cellular metabolic landscapes and mutational motifs caused by exposure to potassium bromate compared to other redox agents.

The most striking change induced by potassium bromate, which is used in breadmaking in the U.S., was a dramatic depletion of an antioxidant called glutathione. In addition, exposure to potassium bromate led to increased levels of the redox agent superoxide. The importance of the discovery of a mutational signature for potassium bromate is underscored by the finding of its increased mutational load in human cancer samples. According to the authors, this study may provide the framework for identifying biomarkers of different types of redox stress in human tumors. [Read related article.]

Citation: Degtyareva NP, Placentra VC, Gabel SA, Klimczak LJ, Gordenin DA, Wagner BA, Buettner GR, Mueller GA, Smirnova TI, Doetsch PW. 2023. Changes in metabolic landscapes shape divergent but distinct mutational signatures and cytotoxic consequences of redox stress. Nucleic Acids Res; doi: 10.1093/nar/gkad305. [Online 20 April 2023]. (JW)

Tracking trends in phthalate and phenol exposure during pregnancy

Pregnant women’s exposure to hormone-disrupting chemicals called phthalates and phenols have decreased over time, while exposure to phthalate replacements has increased, according to NIEHS researchers and their collaborators.

Phthalates and phenols are ubiquitous in the environment due to their widespread use in personal care products, food packaging, household items, and building materials. While these chemicals pose serious health risks for the general population, they are of particular concern for pregnant women. Exposure to many phthalates and phenols is declining as replacement chemicals are introduced. Yet there is little information on temporal trends or predictors of exposure to these newer compounds, especially among pregnant populations.

To address this knowledge gap, the researchers measured concentrations of 12 phthalate metabolites, four phthalate replacement metabolites, and 12 phenols in urine samples collected from 900 pregnant women between 2007 and 2018. Biomarker concentrations of all phthalate metabolites and most phenols declined over time. On the other hand, there was a sharp increase in metabolite concentrations of phthalate replacements, such as di(isononyl) cyclohexane-1,2-dicarboxylic acid, diisononyl ester (DINCH), and di-2-ethylhexyl terephthalate (DEHTP).

In addition, biomarker concentrations of most chemical exposures were highest among non-Hispanic Black and Hispanic participants. According to the researchers, these findings highlight the importance of continued study of these new and emerging chemicals of concern.

Citation: Bommarito PA, Stevens DR, Welch BM, Weller D, Meeker JD, Cantonwine DE, McElrath TF, Ferguson KK. 2023. Temporal trends and predictors of phthalate, phthalate replacement, and phenol biomarkers in the LIFECODES Fetal Growth Study. Environ Int 174:107898. (JW)

Methylation markers of biological age may capture hypertension risk

Higher biological age, as evidenced by changes in DNA methylation, is associated with a woman’s risk of developing hypertension, according to NIEHS researchers.

Hypertension, or high blood pressure, is common among older individuals, and leads to risk of cardiovascular diseases. DNA methylation (DNAm) — a chemical change where methyl groups are attached to particular locations within DNA that are associated with gene transcription — can be used as a clock to estimate biological age and risk of age-related disease. However, few studies have explored biological age metrics, such as DNAm, in relation to hypertension.

The NIEHS researchers used three DNAm metrics to examine biological age associations with hypertension prevalence and incidence. Blood DNAm profiles were generated for 4,419 women, along with data on hypertension at enrollment and during 10 years of annual follow-up.

They noted that methylation-based biological age estimates exceeded a woman’s chronological age before the onset of hypertension and appeared to remain elevated in the years after diagnosis and treatment. The findings help clarify the links between age-related molecular changes and hypertension and suggest that biological age metrics could be useful for hypertension risk stratification.

Citation: Kresovich JK, Sandler DP, Taylor JA. 2023. Methylation-based biological age and hypertension prevalence and incidence. Hypertension 80(6):1213–1222. (MD)

Curbing coronaviruses by delaying viral replication

A pair of mammalian proteins called TUT4/7 marks viral RNAs for decay and delays the replication of coronaviruses, according to NIEHS researchers and their collaborators.

Identifying factors that regulate viral RNA processing is critical for developing novel treatments. One intriguing phenomenon is that the viral poly(A) tails — stretches of RNA that consist only of adenine bases — change in length during infection, thereby potentially contributing to viral replication.

To investigate the underlying mechanisms, the researchers focused on a biochemical process called terminal uridylation, which is the addition of a nucleobase called uracil to the end of the RNA. They studied this process by infecting cells with a coronavirus called mouse hepatitis virus.

The experiments revealed that coronaviruses’ poly(A) tails show two types of uridylation, one of which is dependent on mammalian proteins called terminal uridylyl-transferase 4 (TUT4) and terminal uridylyl-transferase 7 (TUT7), collectively referred to as TUT4/7. Depletion of these proteins reduced the levels of uridylation of short tails, promoted viral replication, and increased the amount of viral RNA.

Based on the findings, the researchers propose that TUT4/7 directly targets RNAs to promote their decay, which, in turn, limits the replication capacity of the virus. Taken together, the results reveal a previously unexplored pathway in the RNA processing of coronaviruses.

Citation: Gupta A, Li Y, Chen SH, Papas BN, Martin NP, Morgan M. 2023. TUT4/7-mediated uridylation of a coronavirus subgenomic RNAs delays viral replication. Commun Biol 6(1):438. (JW)