Structurally distinct polycyclic aromatic hydrocarbons induce differential transcriptional responses in developing zebrafish

Abstract

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous in the environment as components of fossil fuels and by-products of combustion. These multi-ring chemicals differentially activate the Aryl Hydrocarbon Receptor (AHR) in a structurally dependent manner, and induce toxicity via both AHR-dependent and-independent mechanisms. PAH exposure is known to induce developmental malformations in zebrafish embryos, and recent studies have shown cardiac toxicity induced by compounds with low AHR affinity. Unraveling the potentially diverse molecular mechanisms of PAH toxicity is essential for understanding the hazard posed by complex PAH mixtures present in the environment. We analyzed transcriptional responses to PAH exposure in zebrafish embryos exposed to benz(a)anthracene (BAA), dibenzothiophene (DBT) and pyrene (PYR) at concentrations that induced developmental malformations by 120 hours post-fertilization (hpf). Whole genome microarray analysis of mRNA expression at 24 and 48 hpf identified genes that were differentially regulated over time and in response to the three PAH structures. PAH body burdens were analyzed at both time points using GC-MS, and demonstrated differences in PAH uptake into the embryos. This was important for discerning dose-related differences from those that represented unique molecular mechanisms. While BAA misregulated the least number of transcripts, it caused strong induction of cyp1a and other genes known to be downstream of the AHR, which were not induced by the other two PAHs. Analysis of functional roles of misregulated genes and their predicted regulatory transcription factors also distinguished the BAA response from regulatory networks disrupted by DBT and PYR exposure. These results indicate that systems approaches can be used to classify the toxicity of PAHs based on the networks perturbed following exposure, and may provide a path for unraveling the toxicity of complex PAH mixtures.

Retene causes multifunctional transcriptomic changes in the heart of rainbow trout ( Oncorhynchus mykiss ) embryos

“…Though the highest mRNA levels were observed on day 1, the induction was sustained throughout the time course. The high up-regulation of cyp1 genes is in line with previous research (Table S14) (Carney et al, 2006,Goodale et al, 2013,Scott et al, 2011. Also other members of the AhR gene battery were up-regulated (Table S13).…”

“…The ahr gene family is well represented in this group: Several ahr2s were up-regulated on day 1, ahr2a on day 3 as well, and ahr repressor (ahrr) at every time point. The result is in line with previous research showing that ahr2 and ahrr are up-regulated in fish early life stages exposed to AhR agonists (Carney et al, 2006,Goodale et al, 2013. Rainbow trout AHR2α and AHR2β and all members of Atlantic salmon AHR2 protein family are capable of binding TCDD, and may be involved in AhR signaling (Pollenz et al, 2002,Hansson andHahn, 2008).…”

“…AhR-ARNT-dependent transcriptional regulation is needed for toxic effects to take place, but the downstream targets are not well resolved (Antkiewicz et al, 2006,Carney et al, 2004,Prasch et al, 2003,Prasch et al, 2006. Some information has been gained from studies on zebrafish embryos exposed to TCDD, but only a little is known about the effects of AhR-agonistic PAHs (Carney et al, 2006,Chen et al, 2008,Goodale et al, 2013. Furthermore, the molecular mechanisms have to our knowledge not been studied in depth in salmonids at all.…”

Ligand Promiscuity of Aryl Hydrocarbon Receptor Agonists and Antagonists Revealed by Site-Directed Mutagenesis

“…This idea is supported by gene microarray studies which have reported the ability of different AhR ligands to not only induce expression of a common set of gene products but to also induce a distinctly different, ligandspecific set of AhR-dependent gene products (21)(22)(23). Ligandspecific modulation of the AhR signaling pathway has been suggested to result from ligand-dependent changes in the AhR which could allow it to interact with different nuclear factors or dimerization partners, to bind to unconventional DREs or unique DNA sequences, and/or to be bound by different coactivators (24)(25)(26)(27).…”

WFIKKN1 and WFIKKN2: “Companion” proteins regulating TGFB activity

“…Regarding WFIKKN1, it has been reported to be overexpressed in response to various compounds (Table 3) such as polycyclic aromatic hydrocarbons in zebrafish [54], retinoic acid in human embryonic stem cells [55], the caspase inhibitor RBC1023 [56] in mice and the gastric inhibitory polypeptide in rats [57]. The WFIKKN1 gene is also downregulated in mouse embryonic stem cells overexpressing the N-terminal domain of the Dnmt3a1 DNA methyltransferase probably via epigenetic silencing [58].…”