Editing the insulin-like growth factor binding protein-2b gene in rainbow trout

Authors: Cleveland, Beth; YAMAGUCHI, GINNOSUKE - Hokkaido University; Radler, Lisa; SHIMIZU, MUNETAKA - Hokkaido University

Submitted to: International Congress on the Biology of Fish
Publication Type: Abstract Only
Publication Acceptance Date: 3/25/2018
Publication Date: 7/15/2018
Citation: Cleveland, B.M., Yamaguchi, G., Radler, L.M., Shimizu, M. 2018. Editing the insulin-like growth factor binding protein-2b gene in rainbow trout [abstract]. International Congress on the Biology of Fish. P-165-E.

Interpretive Summary:

Technical Abstract: In salmonids, the majority of circulating insulin-like growth factor-I (IGFI) is bound to IGF binding proteins (IGFBP), with IGFBP2b being the most abundant IGFBP in circulation. In this study we used CRISPR/Cas9 methodology to disrupt expression of a functional IGFBP2b protein. Twenty four rainbow trout were produced with mutations in the IGFBP2b1 and IGFBP2b2 genes and gene variants were characterized by next-generation sequencing. The percentage of gene mutagenesis was variable; between 5 – 88% of IGFBP2b variants amplified from individual fish differed from the reference gene sequences; those predicted to cause frame-shift mutations in individual fish were between 2 – 69% of IGFBP2b variants. Analysis of IGFBP2b protein via ligand binding assay indicated between 4 – 93% reduction in plasma IGFBP2b abundance compared to wild type fish; reductions in individual fish were positively correlated with percent gene mutagenesis (P=0.002, R2=0.47). These findings indicate that gene editing is useful to unravel IGF-related endocrine mechanisms in rainbow trout.