Transcriptional changes associated with domoic acid (DomA) exposure at 2 d postfertilization (dpf). (A) Experimental design. Tanks of three adult fish (2 females, 1 male) of Tg(mbp:EGFP-CAAX) background were bred and the embryos exposed to DomA (0.14 ng) or vehicle at 2 dpf. Pools of six embryos within a given treatment from each tank were then sampled at 3 and 7 dpf for RNA sequencing. Three pools per treatment represented the three biological replicates. For functional analyses, myelin sheath labeling was assessed at 5 dpf and startle response was assessed at 7 dpf prior to RNA sequencing. The results confirming differences in behavior and myelin labeling between DomA-exposed fish and controls used for RNA sequencing are shown in Figure S6. (B) A multidimensional scaling plot shows clustering of samples based on overall differences in expression profiles. (C,D) Mean-difference plots compare the log fold changes of genes in DomA-exposed vs. control fish at the 3 dpf- and 7 dpf-sampling times. Hollow teal circles (+1) represent genes that were significantly up-regulated in DomA-exposed fish relative to their controls, whereas filled magenta circles (–1) represent genes that were significantly down-regulated in DomA-exposed fish relative to their controls. Significance was determined with a genewise negative bionomial generalized linear model with a quasi-likelihood test. p-Values were adjusted for using a 5% false discovery rate cutoff. Tables S30 and S31 contain the results of the functional enrichment analysis done using the differentially expressed genes shown in (C). Excel Table S1 contains the list of genes that were differentially expressed in DomA-exposed fish at 3 dpf, and Excel Table S2 contains the list of genes that were differentially expressed in DomA-exposed fish at 7 dpf. Note: CPM, counts per million; FC, fold change.
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