Insulin-like growth factor binding protein-2 (IGFBP-2) in Japanese flounder, Paralichthys olivaceus: molecular cloning, expression patterns and hormonal regulation during metamorphosis

Abstract

In this study, we cloned and characterized cDNA sequences of two insulin-like growth factor binding protein-2 (IGFBP-2a and IGFBP-2b) from Japanese flounder, Paralichthys olivaceus. The full-length cDNA of IGFBP-2a is 1,046bp long and consists an open frame (ORF) of 876bp, a 5'-untranslated region (UTR) of 125bp and a 3'-UTR of 45bp. IGFBP-2b is 1,067bp, including a 5'-UTR of 53bp, a 3'-UTR of 198bp and an ORF of 816bp. Real-time quantitative PCR results revealed that IGFBP-2a -2b mRNA was expressed in all detected tissues. Interestingly, the levels of IGFBP-2a mRNA in all detected tissues were higher in female than male, but IGFBP-2b was precisely the opposite. At different embryonic stages, the levels of IGFBP-2a mRNA were typically higher than IGFBP-2b. After hatching, IGFBP-2a mRNA was gradually decreased to a relatively lower level. However, the expression of IGFBP-2b mRNA was increased after hatching, including 3, 7, 10, 14, 17, 20 and 23days post-hatching (dph), and it presents a higher level until 29 (metamorphic climax), 36 (post-climax) and 41dph (the end of metamorphosis). In levothyroxine sodium salt (T4, the main form of thyroid hormone in animals)-treated and thiourea (TU)-treated larvae, the expressions of IGFBP-2a had not visibly changed, except in T4-treated 17dph larvae. The expressions of IGFBP-2b mRNA were distinctly increased from 17 to 23dph, but suddenly dropped to a lower level in and after 29dph. However, the levels of IGFBP-2b mRNA during metamorphosis were greatly down-regulated after TU treatment. These results provided basic information for further studies on the role of IGF system in flatfish development and metamorphosis.