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Papers of the Month

Intramural
By Rachel Carroll, Douglas Ganini da Silva, Kelly Shipkowski, Heather Vellers, and Qing Xu

NTP screens for genotoxicity with high-throughput techniques

National Toxicology Program (NTP) researchers and collaborators evaluated the usefulness of quantitative high-throughput approaches to assess the genotoxicity, or the ability to cause DNA damage, of compounds in the Tox21 library of 10,000 compounds. To do this, they assessed p53 activity in human colon cancer cells (HCT-116). p53 is a transcription factor that is activated in response to cellular stress, particularly DNA damage.

The Tox21 high-throughput screening program is part of a collaborative effort to institute new methods for toxicity testing. Standard toxicology tests cannot keep pace with the number of new and existing chemicals used in the United States. The goal of the Tox21 program is to develop rapid, predictive, mechanism-based methods to broaden the coverage of compounds with toxicological characterization using a library of 10,000 diverse compounds. To date, screening has focused on nuclear receptor and stress response pathway signaling. The p53 assay falls into the second category.

Results showed that 4.7 percent of compounds activated p53. Additional data mining revealed significant associations between these compounds and elevated expression of p53 downstream genes. Further analysis revealed specific associations between p53 activation and specific chemical substructures. The authors concluded that assessment of p53 activation was useful for characterization of genotoxic potential. (KS)

Citation: Witt KL, Hsieh JH, Smith-Roe SL, Xia M, Huang R, Zhao J, Auerbach SS, Hur J, Tice RR. 2017. Assessment of the DNA damaging potential of environmental chemicals using a quantitative high-throughput screening approach to measure p53 activation. Environ Mol Mutagen 58(7):494–507.

Season of conception predicts preeclampsia in Norway

A team led by NIEHS researchers studied births in Norway and found a seasonal pattern in the occurrence of preeclampsia, a dangerous pregnancy complication that usually presents with high blood pressure and protein in urine. Mothers who conceived during the month of May had the highest rates of preeclampsia.Mothers who conceived in November had the lowest rates. The findings offer important evidence that a seasonally varying environmental factor contributes to the risk of preeclampsia.

With advanced statistical modeling, the researchers used harmonic Cox regression to model confounder-adjusted effects of the estimated day of the year of conception on risk of preeclampsia. They applied the model to 356,662 births from the Medical Birth Registry of Norway between 1999 and 2009.

Regardless of parity, the region of Norway, fetal sex, and maternal smoking status, the same seasonal pattern of preeclampsia risk appeared. With a risk amplitude of approximately 20 percent, the highest rates occurred in spring conception and the lowest in autumn conceptions. Taken together, these results suggest there is a seasonal driver for preeclampsia, with effects that are not modified by parity, region, fetal sex, or smoking. (HV)

Citation: Weinberg CR, Shi M, Basso O, DeRoo LA, Harmon Q, Wilcox AJ, Skjaerven R. 2017. Season of conception, smoking, and preeclampsia in Norway. Environ Health Perspect 125(6):067022.

Pet allergies associated with increased asthma morbidity

Exposure to higher levels of allergens from dogs, cats, or both, increased numbers of asthma attacks among individuals with asthma and sensitivities to these allergens, according to NIEHS scientists and their collaborators. Although exposure to dogs and cats can increase asthma symptoms among pet-sensitive individuals, until now, no studies investigated the impact of residential dog and cat allergen exposures on asthma morbidity at the national level.

The researchers analyzed nationally representative data from the National Health and Nutrition Examination Survey. The survey assessed levels of dog and cat allergens in dust collected from participants’ homes, drew blood to test sensitivity to dogs and cats, and queried participants about asthma.

The data estimated that among the U.S. population, dog allergen exposure contributed to 1,161,000 excess asthma attacks among dog-sensitive asthmatics, whereas cat exposure attributed to 549,000 excess attacks among cat-sensitive individuals. The results underscore the importance of reducing pet allergen levels in homes of pet-sensitive asthmatics. Pet allergen reduction could have major impacts on asthma morbidity and health care use. (RC)

Citation: Gergen PJ, Mitchell HE, Calatroni A, Sever ML, Cohn RD, Salo PM, Thorne PS, Zeldin DC. 2017. Sensitization and exposure to pets: the effect on asthma morbidity in the US population. J Allergy Clin Immunol Pract; doi: 10.1016/j.jaip.2017.05.019 [Online 7 July 2017].

Las1 coordinates with Grc3 to cleave RNA

Researchers from NIEHS and the National Cancer Institute deciphered the molecular mechanisms responsible for the activity of Las1, a RNA-cutting enzyme essential for cell viability and generation of ribosomes. The findings offer new insights into the development of protein-guided gene editing tools and of neurological diseases linked to Las1 mutations.

As the factory for protein synthesis in cells, ribosomes are composed of RNAs and associated proteins. The ribosomal RNAs (rRNAs) come from the precursor rRNAs, pre-rRNAs, with spacer sequences that need to be removed by nucleases, including Las1. The sites cleaved by Las1 are phosphorylated by the polynucleotide kinase Grc3, which signals other enzymes to further process the pre-rRNAs. Las1 was shown to interact with Grc3 for ribosome assembly, but whether the interaction is required for their enzymatic activities was unclear.

Researchers in this study observed that Las1 associated with Grc3 and two Las1/Grc3 heterodimers formed a superdimer. Las1 had weak activity, but displayed efficient cleavage in the presence of Grc3. The two proteins relied on each other for functions and recognition of the cutting sites. The study also identified protein domains that were indispensable for the formation of superdimer and enzymatic activities, establishing a working model for Las1/Grc3-mediated pre-rRNA processing. (QX)

Citation: Pillon MC, Sobhany M, Borgnia MJ, Williams JG, Stanley RE. 2017. Grc3 programs the essential endoribonuclease Las1 for specific RNA cleavage. Proc Natl Acad Sci U S A 114(28):E5530−E5538.

BRG1 is required for development of the lymphatic system

NIEHS scientists demonstrated the central importance of the chromatin-remodeling protein Brahma-related gene 1 (BRG1) for the development of the lymphatic system during embryo development. The research could potentially lead to novel therapies for circulatory disorders, cancer metastases, and other illnesses.

The team found that when BRG1 was inactivated late in the development of mice, the embryos were not viable and presented morphological defects in the lymphatic and cardiovascular systems. Using genome-wide RNA sequencing, microarray analyses, and quantitative real-time polymerase chain reaction, the researchers found lower levels of several lymphatic vessel markers in the embryos. Other genes that support the survival of lymphoid cells were also decreased.

Analysis of human cells in culture showed that increased expression of BRG1 resulted in a substantial increase in the levels of specific lymphatic vessel markers. However, when BRG1 was silenced, the cells in culture exhibited less ability to form vessel structures. The authors also determined that BRG1 specifically binds to promoter regions of key regulators known to promote the development of the lymphatic system in embryos. This work advances the understanding of lymphatic gene dysregulation in lymphedema and other conditions. (DGS)

Citation: Singh AP, Foley J, Tandon A, Phadke D, Kinyamu HK, Archer TK. 2017. A role for BRG1 in the regulation of genes required for development of the lymphatic system. Oncotarget 8:54925–54938.

(Rachel Carroll, Ph.D., is a research fellow in the NIEHS Biostatistics and Computational Biology Branch. Douglas Ganini da Silva, Ph.D., is a research fellow in the NIEHS Free Radical Metabolism Group. Kelly Shipkowski, Ph.D., is an Intramural Research and Training Award (IRTA) fellow in the NTP Systems Toxicology Group. Heather Vellers, Ph.D., is an IRTA fellow in the NIEHS Environmental Genetics Group. Qing Xu is a biologist in the NIEHS Metabolism, Genes, and Environment Group.)