Developmental expression and regulation of flavin-containing monooxygenase by the unfolded protein response in Japanese medaka (Oryzias latipes)

Abstract

Flavin-containing monooxygenases (FMOs) play a key role in xenobiotic metabolism, are regulated by environmental conditions, and are differentially regulated during mammalian development. Japanese medaka (Oryzias latipes) are a common model organism for toxicological studies. The goal of the current research was to characterize developmental expression and regulation of FMOs in Japanese medaka embryos to better understand the role of FMOs in this model species. Five putative medaka fmos were characterized from the medaka genome through the National Center for Biotechnology Information (NCBI) database by protein motifs and alignments, then identified as fmo4, fmo5A, fmo5B, fmo5C and fmo5D for the current study. Fmo gene expression was analyzed at 1dpf, 3dpf, 6dpf and 9dpf and distinct developmental patterns of expression were observed. Fmo4 and fmo5D increased 3-fold during mid organogenesis (6dpf), while fmo5B and fmo5C decreased significantly in early organogenesis (3dpf) and fmo5A was unaltered. Promoter analysis was performed for transcription factor binding sites and indicated regulation by developmental factors and a role for the unfolded protein response in fmo modulation. Fmo regulation by the UPR was assessed with treatments of 1μg/ml, 2μg/ml, and 4μg/ml Tunicamycin (Tm), and 2mM and 4mM dithiothreitol (DTT), well-known inducers of endoplasmic reticulum stress, for 24h from 5–6dpf. High concentrations to Tm induced fmo4 and fmo5A up to two-fold, while DTT significantly decreased expression of fmo5A, fmo5B, and fmo5C. Results suggest that medaka fmos are variably regulated by the UPR during organogenesis with variable developmental expression, and suggesting potential stage-dependent activation or detoxification of xenobiotics.