Molecular and functional characterization of two distinct IGF binding protein-6 genes in zebrafish

Insulin-like growth factor binding proteins (IGFBPs) are high-affinity binding partners for IGFs and play important roles in modulating IGF activities. In this study, we have identified and characterized two functional IGFBP-6 genes in zebrafish. Structural, phylogenetic, and comparative genomic analyses indicate that they are co-orthologs of the human IGFBP-6 gene. To gain insight into how the duplicated genes may have evolved through partitioning of ancestral functions, gene expression and functional studies were carried out. In adult fish, IGFBP-6a mRNA was most abundantly expressed in the muscle. The levels of IGFBP-6a mRNA in nonmuscle tissues were very low or barely detectable. In comparison, the levels of IGFBP-6b mRNA were high in the brain, heart, and muscle, but very low or undetectable in other adult tissues. During embryogenesis, the IGFBP-6a mRNA levels were relatively low. The IGFBP-6b mRNA levels were low during the initial 48 h. They became significantly higher at 72 and 96 h postfertilization. Overexpression of zebrafish IGFBP-6a and IGFBP-6b caused a similar degree of reduction in body size and developmental rate. No notable effects were observed on cell fate or patterning in these transgenic fish. These data suggest that the duplicated igfbp-6 genes encode two functionally similar proteins, but they have evolved distinct spatial and temporal expression patterns. These findings are consistent with the notion of an additional gene duplication event in teleost fish and have provided novel insight into the structural and functional evolution of the IGFBP gene family.