Transcriptome Alterations Following Developmental Atrazine Exposure in Zebrafish Are Associated with Disruption of Neuroendocrine and Reproductive System Function, Cell Cycle, and Carcinogenesis

Abstract

Atrazine, a herbicide commonly applied to agricultural areas and a common contaminant of potable water supplies, is implicated as an endocrine-disrupting chemical (EDC) and potential carcinogen. Studies show that EDCs can cause irreversible changes in tissue formation, decreased reproductive potential, obesity, and cancer. The U.S. Environmental Protection Agency considers an atrazine concentration of ≤ 3 ppb in drinking water safe for consumption. The specific adverse human health effects associated with a developmental atrazine exposure and the underlying genetic mechanisms of these effects are not well defined. In this study, zebrafish embryos were exposed to a range of atrazine concentrations to establish toxicity. Morphological, transcriptomic, and protein alterations were then assessed at 72h postfertilization following developmental atrazine exposure at 0, 0.3, 3, or 30 ppb. A significant increase in head length was observed in all three atrazine treatments. Transcriptomic profiles revealed 21, 62, and 64 genes with altered expression in the 0.3, 3, and 30 ppb atrazine treatments, respectively. Altered genes were associated with neuroendocrine and reproductive system development, function, and disease; cell cycle control; and carcinogenesis. There was a significant overlap (42 genes) between the 3 and 30 ppb differentially expressed gene lists, with two of these genes (CYP17A1 and SAMHD1) present in all three atrazine treatments. Increased transcript levels were translated to significant upregulation in protein expression. Overall, this study identifies genetic and molecular targets altered in response to a developmental atrazine exposure to further define the biological pathways and mechanisms of toxicity.

Water Contaminated With the Herbicide Atrazine: Alteration of Cancer Associated Tumor Protein D52-like 1 (tpd52l1) Gene Using the Zebrafish Model

“…A previous study in the Freeman laboratory provided insight into atrazine exposure during embryogenesis of the zebrafish (Weber et al, 2013). The study encompassed the developmental time points of 1 to 72 hours postfertilization (hpf; the end of embryogenesis) and exposure of atrazine at various concentrations (0, 0.3, 3, or 30 ppb) to understand the effects of atrazine during development.…”

“…The transcriptomic study revealed alterations in expression in a couple hundred genes, some of which have known roles in carcinogenesis. Of these genes, expression studies through (quantitative polymerase chain reaction) qPCR and western blot were conducted and cyp17a1, glo1, sik2, and tpd52l1 displayed significant upregulation in gene and protein expression at 72 hpf (Weber et al, 2013). The role of these genes goes beyond carcinogenesis; they have shown connections to cell cycle regulation, cell signaling, and reproduction.…”

“…These test concentrations represent 0.1 to 10 times the current regulated concentration in drinking water in the US. These concentrations were prepared using a stock solution of technical grade atrazine (98.1% purity) (CAS 1912-24-9; ChemService, West Chester, PA) and were prepared as described in prior studies (Weber et al, 2013;Wirbisky et al, 2015). Following exposure to the atrazine treatment, the larvae were rinsed with system water as previously described for experimental use (Weber et al, 2013).…”

“…First, information was gathered and collectively recorded to include any roles the genes, identified with altered expression from Weber et al (2013), play in carcinogenesis. From this analysis, tpd52l1 was chosen as the target gene for this study.…”

“…Quantification of gene expression was conducted using established protocols to obtain developmental time point data for tpd52l1 (Weber et al, 2013). To do this transcript analysis for all biological repetitions, 50 embryos were collected into each petri dish, comprising a single replicate.…”