Evaluating the impact of chemical mixtures
Some cell-based assays may be better than others at capturing the combined effects of individual chemicals, according to researchers from the Division of Translational Toxicology.
The goal of the U.S. Tox21 program is to develop high-throughput screening approaches for toxicity testing. In the new study, the researchers examined how effectively the screening approaches predicted responses to chemical mixtures. They used two estrogen receptor alpha (ER-alpha) agonist assays: ER-bla and ER-luc. These cell-based assays, which measure the extent to which chemicals activate estrogen signaling pathways, are crucial for assessing the potential for environmental chemicals to disrupt the endocrine system.
Models that predicted the joint impact of mixtures based on data from individual chemicals tended to overestimate the responses compared with data from real mixtures in the ER-bla assay. In contrast, model predictions resembled real responses in the ER-luc assay, which is considered more sensitive at detecting ER-alpha activation. The results suggest that the ER-luc assay is preferred for the high-throughput analysis of mixtures.
Based on the findings, the researchers also suggest that individual chemicals and mixtures should be analyzed using the same experimental setup. They also presented two methods for estimating missing data to improve estimations of mixture effects. According to the authors, future efforts should explain discrepancies between modeled predictions and observed responses to improve models and testing approaches. (JW)
Citation: Parham F, Eccles KM, Rider CV, Sakamuru S, Xia M, Huang R, Tice RR, Dinse GE, DeVito MJ. 2025. Lessons learned from evaluating defined chemical mixtures in a high-throughput estrogen receptor assay system. Toxicol Sci 205(1):191-204.
Changes in mitochondrial function may affect gene regulation
Exposure to some environmental chemicals may alter a cell’s mitochondrial inner membrane potential (ΔΨm), according to researchers from the Division of Translational Toxicology.
Mitochondria are membrane-bound cell organelles that generate most of the chemical energy needed to power the cell's biochemical reactions. ΔΨm is a crucial electrochemical gradient across the inner mitochondrial membrane; it is essential for energy production and various mitochondrial functions. However, the effects of increases in the ΔΨm, known as hyperpolarization, have not been clear, although higher values have been linked to lung disease, brain cancer, and ovarian cancer.
To fill this knowledge gap, the researchers used genetically modified cells to study the impact of high resting levels of ΔΨm. The results revealed long-lasting molecular, metabolic, and genomic effects of mitochondrial hyperpolarization. Because of changes in phospholipids — an essential membrane component — there was an increase in nuclear DNA methylation, affecting the activity of mitochondrial, carbohydrate, and lipid genes.
Additional results showed that these effects can be mimicked by exposing cells to certain environmental chemicals, such as food additives and medications. Other compounds that tested positive in inducing ΔΨm hyperpolarization in a Tox21 dataset, such as pesticides and anti-fouling agents, might also recapitulate the same phenotype. According to the authors, ΔΨm could potentially be targeted to better understand disease states and improve human health. (JW) (See related article.)
Citation: Mori MP, Lozoya OA, Brooks AM, Bortner CD, Nadalutti CA, Ryback B, Rickard BP, Overchuk M, Rizvi I, Rogasevskaia T, Huang KT, Hasan P, Hajnóczky G, Santos JH. 2025. Mitochondrial membrane hyperpolarization modulates nuclear DNA methylation and gene expression through phospholipid remodeling. Nat Commun 16(1):4029.
Steric gate helps DNA ligase 1 differentiate between DNA and RNA
NIEHS researchers and their collaborators have discovered how the enzyme DNA ligase 1 (LIG1) prevents mistakes when sealing broken DNA strands.
DNA ligation is the process of joining two DNA fragments to form a single strand. The multistep ligation reaction is catalyzed by the enzyme LIG1, which finalizes DNA replication in the nucleus by accurately sealing nicks. DNA ligases are highly specific and prefer DNA’s deoxyribose sugar molecule over RNA’s ribose sugar molecule, making sure that RNA does not mistakenly get included in DNA strands. However, it is not clear how LIG1 differentiates between ribonucleotides and deoxyribonucleotides.
To investigate, the researchers studied the crystal structures of LIG1 and LIG1 mutants when interacting with DNA and RNA. By solving these structures and through various biochemical approaches, the researchers found that LIG1 uses a conversed aromatic amino acid F872 as a steric gate, physically blocking RNA from being incorporated. When they mutated this gate, LIG1 lost much of its ability to distinguish between DNA and RNA, leading to incorrect ligation.
These findings demonstrate how LIG1 maintains DNA accuracy and stability. According to the authors, DNA ligases are critical for faithful replication of the eukaryotic genome and may use similar strategies to exclude ribonucleotide incorporation as high-fidelity DNA polymerases. (SS)
Citation: Tumbale PP, Jurkiw TJ, Krahn JM, Bokil NV, Admiraal SJ, Pedersen LC, Williams JS, Kunkel TA, O'Brien PJ, Williams RS. 2025. Molecular basis for RNA discrimination by human DNA ligase 1. Nucleic Acids Res 53(7):gkaf299.
Phthalate exposure in early pregnancy may be linked to preterm birth
Exposure to phthalates during the first two trimesters of pregnancy may increase the risk of preterm birth, according to NIEHS researchers and their collaborators.
Phthalates are chemicals commonly present in many everyday products, such as personal care items, packaging materials, and medical supplies. Previous studies have reported that prenatal phthalate exposure may result in adverse pregnancy outcomes, including preterm birth.
To determine critical periods of susceptibility between conception and delivery, the researchers examined urinary concentrations of phthalate metabolites in 6,045 participants. Consistent with their previous analysis, they observed associations between several phthalate metabolites and preterm birth. In addition, for di-2-ethylhexyl phthalate metabolites, they observed that the association with preterm birth was dependent on the timing of exposure, and effects were greatest for phthalate exposure measurements in the first and second trimester.
These findings indicate that phthalate exposure during the early stages of pregnancy may increase the risk of preterm birth. According to the authors, future studies to evaluate critical periods of susceptibility are needed. (SS)
Citation: Friedman A, Welch BM, Keil AP, Bloom MS, Braun JM, Buckley JP, Dabelea D, Factor-Litvak P, Meeker JD, Michels KB, Padmanabhan V, Starling AP, Weinberg CR, Aalborg J, Alshawabkeh AN, Barrett ES, Binder AM, Bradman A, Bush NR, Calafat AM, Cantonwine DE, Christenbury KE, Cordero JF, Engel SM, Eskenazi B, Harley KG, Hauser R, Herbstman JB, Holland N, James-Todd T, Jukic AMZ, Lanphear BP, McElrath TF, Messerlian C, Newman RB, Nguyen RHN, O'Brien KM, Rauh VA, Redmon JB, Rich DQ, Rosen EM, Sathyanarayana S, Schmidt RJ, Sparks AE, Swan SH, Wang C, Watkins DJ, Weinberger B, Wenzel AG, Wilcox AJ, Yolton K, Zhang Y, Zota AR, Ferguson KK. 2025. Periods of susceptibility for associations between phthalate exposure and preterm birth: results from a pooled analysis of 16 US cohorts. Environ Int 198:109392.
Residential natural hazard risk may put people at risk for cardiovascular disease-related outcomes
Living in a disaster-prone area is linked to higher prevalence of risk factors related to cardiovascular disease, according to NIEHS researchers and their collaborators.
Adverse health outcomes have been documented following natural disasters, such as floods, wildfires, and drought. Yet the consequences of living in places with an increased risk of frequent natural hazards have not been well studied. To better understand these risks, the researchers studied the link between living in residential areas prone to natural disasters and the prevalence of diabetes, high blood pressure, and obesity, which are risk factors related to cardiovascular disease.
The researchers analyzed data from 32,608 adults who participated in the Gulf Long-term Follow-up Study, a large prospective cohort of oil spill cleanup workers and nonworkers enrolled following the 2010 Deepwater Horizon disaster. Data included interview-based questionnaires and home visits for a subset of participants. Researchers linked the National Risk Index, which quantifies hazard risk at the census-tract level, to participants’ geocoded residential addresses. The index quantifies risk scores overall for all hazards, as well as individual event types, including floods, heatwaves, and hurricanes.
The results showed that the overall hazard risk was associated with higher high blood pressure. All three cardiovascular risk factors were more prevalent when heatwave risk was elevated, and the prevalence of diabetes was higher for those living in areas with higher hurricane risk. According to the authors, the findings broaden the literature on the negative impact of natural hazards on human health. (JW)
Citation: Lawrence KG, Sweeney MR, Werder EJ, Zuzak C, Gall M, Emrich CT, Buller ID, Jackson WB 2nd, Chen D, Christenbury KE, Engel LS, Sandler DP. 2025. Prevalence of cardiovascular disease risk factors associated with residential natural hazard risk. Sci Total Environ 977:179335.
Calcium signals synchronized to coordinate activities inside cells
Live cell imaging shows how the timing of calcium flow is coordinated across the cell, potentially enabling the simultaneous activation of key signaling molecules, according to NIEHS researchers and their collaborators.
Ca2+ ions, or calcium ions, are essential for various biological processes. They are used inside almost all cell types across a variety of organisms. Ca2+ oscillations convey important information and activate responses related to cell metabolism, growth, and death. Ca2+ enters cells through Ca2+ release-activated Ca2+ (CRAC) channels in the plasma membrane. But the role of CRAC channels in Ca2+ oscillations has not been clear.
One model proposes that the channels stay open throughout an oscillation, providing a continuous flux of Ca2+, whereas another model suggests that CRAC channels activate and deactivate during the oscillatory cycle, leading to bursts of Ca2+ entry into cells. To answer this question, the researchers tethered a genetically encoded Ca2+ indicator to Orai1 — a subunit of the CRAC channel. In support of the second model, the results showed that CRAC channel activity fluctuates in synch with cytosolic Ca2+ oscillations.
According to the authors, the synchrony between Ca2+ oscillations and CRAC channel activity may enable signaling molecules to coordinate their activities across the cytosol and the cell periphery. (JW)
Citation: Lin YP, Scappini E, Mirams G, Tucker CJ, Parekh AB. 2025. CRAC channel activity pulsates during cytosolic Ca2+ oscillations. J Biol Chem 301(6):108519.
(Janelle Weaver, Ph.D., is a contract writer for the NIEHS Office of Communications and Public Liaison. Shruti Somai, Ph.D., is a visiting fellow in the Genome Integrity and Structural Biology Laboratory.)
