Figure 3

Morpholino knockdown of putative zebrafish Il1r1. (A) RT-PCR of cabz, zmp, and actb1 transcripts from uninjected embryos, control morpholino injected embryos, and cabz and zmp morphants. (B) Schematic of the cabz01054965 (cabz) gene showing exons 3-6, forward (F) and reverse (R) primers used for analysis (black arrowheads), the location of the splice donor site morpholino (blue line), normal splicing of wt (black lines), and aberrant splicing with the cabz morpholino (red line) that excludes exon 4 (-ex4). (C) Schematic of the zmp:0000000936 (zmp) gene showing exons 4-8, forward (F) and reverse (R) primers used for analysis (black arrowheads), the location of the splice donor site morpholino (blue line), normal splicing of wt (black lines), and aberrant splicing with the zmp morpholino (red lines) that includes intron 5 (+in5), introduce a cryptic splice sites (css), or excludes exon 5 (-ex5). (D) Diagram of the experimental paradigm used to functionally identify zebrafish Il1r1. A zebrafish model of Il-1β-induced systemic inflammation was generated by breeding transgenic lines ubb:Gal4-EcR and uas:Ilβ^mat with or without the neutrophil reporter line mpx:mCherry. Single-celled embryos were injected with morpholino, inflammation was induced with 1 µM Tebufenozide (Teb) at 1 or 2 dpf, and functional analyses were performed from 3-6 dpf to assess morpholino rescue of Il-1β-induced inflammation.