The tryptophan photoproduct 6-formylindolo[3,2-b]carbazole (FICZ) binds multiple AHRs and induces multiple CYP1 genes via AHR2 in zebrafish

Abstract

The tryptophan photooxidation product 6-formylindolo[3,2-b]carbazole (FICZ) has been proposed as a physiological ligand for the mammalian aryl hydrocarbon receptor (AHR), which it binds with high affinity, inducing expression of cytochrome P450 1A1 (CYP1A1). We investigated whether the response to FICZ is evolutionarily conserved in vertebrates by measuring FICZ binding to two zebrafish AHRs (AHR1B and AHR2) and its ability to induce zebrafish CYP1 genes (CYP1A, CYP1B1, CYP1C1, CYP1C2, and CYP1D1) in vivo. Exposure of zebrafish embryos (48 hours-post-fertilization; hpf) to 10 nM FICZ for 6 hours caused strong induction of CYP1A mRNA and a statistically significant but modest induction of CYP1B1 and CYP1C1. Neither CYP1C2 nor CYP1D1 expression was induced by FICZ under the conditions of dose, time or developmental stage examined here. CYP1A induction was significantly greater after 6 hours than after 12 hours of exposure to FICZ, suggesting a rapid degradation of inducer. The 6-hr EC50 values for induction of CYP1A and CYP1B1 by FICZ were 0.6 and 0.5 nM compared to 72-hr EC50 values of 2.3 and 2.7 nM for PCB126, indicating that in zebrafish embryos FICZ is a more potent inducer than PCB126. FICZ at 10 nM was able to completely displace binding of 2,3,7,8-tetrachloro-1,6[3H]-dibenzo-p-dioxin to in vitro-expressed zebrafish AHR2 and AHR1B. Inhibition of AHR2 translation in zebrafish embryos by an AHR2-specific morpholino antisense oligonucleotide decreased the induction of CYP1A and CYP1B1 by FICZ and by PCB126. Together, these results demonstrate that FICZ is a potent AHR agonist in zebrafish, inducing expression of multiple CYP1 genes largely through AHR2. Evolutionary conservation of the response to FICZ is consistent with a possible role as an endogenous signaling molecule acting through the AHR.

Time-dependent expression and activity of cytochrome P450 1s in early life-stages of the zebrafish (Danio rerio)

“…All genes investigated had a temporal decline in gene expression at 72 hpf in common. A similar decline was found for cyp1a induction after exposure to 6-formylindolo[3,2-b]carbazole for 6 h prior to sampling (Jönsson et al 2009). Zebrafish embryos hatch sporadically during the third day (between 48 and 72 hpf) of development.…”

“…In these studies, a decline in gene expression was not reported. In the present study as well as in the study of Jönsson et al (2009), the embryos were exposed until directly before measurement of gene expression, both showing a decline in gene expression around time of hatch.…”

Induction of long interspersed nucleotide element-1 (L1) retrotransposition by 6-formylindolo[3,2- b ]carbazole (FICZ), a tryptophan photoproduct

“…L1 is conserved in organisms from zebrafish to human (4), and a human L1 homolog of Candida albicans is competent for retrotransposition in Saccharomyces cerevisiae (38). Furthermore, cellular responses to FICZ have been reported in both a Xenopus laevis cell line (39) and zebrafish embryos (40), implying that FICZ can induce L1-RTP in various living organisms. Although further study is required, our current observations suggest the possibility that a member(s) of the bHLH/PAS family is involved in the epigenetic mode of L1-RTP.…”

The Aryl Hydrocarbon Receptor in Barrier Organ Physiology, Immunology, and Toxicology

“…Autoimmunity is driven by Th17 cells and ameliorated or prevented by antigen-specific regulatory T cells. In the Du et al, 2005;Fritsche et al, 2007;Kostyuk et al, 2012;John et al, 2013Smith et al, 2013 Circadian rhythmicity Mukai and Tischkau, 2007 Expression of AhR target genes Wei et al, 2000;Oberg et al, 2005;Sciullo et al, 2008;Nair et al, 2009;Wincent et al, 2009;Lee et al, 2010;Luecke et al, 2010;Mohammadi-Bardbori et al, 2012;Rico-Leo et al, 2013;Sumida et al, 2013Jönsson et al, 2009Laub et al, 2010;Wincent et al, 2012;Odio et al, 2013 Genome rearrangement Okudaira et al, 2010Okudaira et al, , 2013 Immune response Apetoh et al, 2010;Bankoti et al, 2010Quintana et al, 2008Veldhoen et al, 2008;Martin et al, 2009;Monteleone et al, 2011;Jeong et al, 2012;Pauly et al, 2012;Qiu et al, 2012;Sibilano et al, 2012;Duarte et al, 2013;Wheeler et al, 2013;Zhou et al, 2013 Nuclear receptor cross-talk Ekins et al, 2008;Reschly et al, 2008;Bunaciu and Yen, 2013 AhR activation can also occur by factors that block/inhibit CYP1A, leading to exceptionally high FICZ levels because FICZ is no longer degraded. In both scenarios (B) and (C), FICZ homeostastic...…”