Ontogeny, tissue distribution, and hormonal regulation of insulin-like growth factor binding protein-2 (IGFBP-2) in a marine fish, Sparus aurata.
In this study, we have cloned insulin-like growth factor binding protein (IGFBP)-2 from a marine hermaphroditic fish species, the gilthead sea bream (Sparus aurata), and determined its structure, ontogeny, tissue distribution, and hormonal regulation. The sea bream IGFBP-2 precursor consists of 286 amino acids, including a putative signal peptide of 22 residues and a mature protein of 264 residues. The overall sequence of sea bream preIGFBP-2 is 52-39% identical to that of zebrafish, chick, mouse, rat, pig, sheep, bovine, and human preIGFBP-2. The cysteine-rich N- and C-terminal domains, which are believed to be important for IGF binding, show a greater degree of conservation with a sequence identity of 73-56% and 53-50%, respectively. Sea bream IGFBP-2 contains 18 cysteine residues and their alignment is identical to those of other vertebrate IGFBP-2s. The Arg-Gly-Asp (RGD) sequence, present in the C-terminal domain of all known IGFBP-2, is also present in the sea bream. Northern blot analysis of RNA samples extracted from adult liver and developing larvae, revealed two transcripts of about 1.3 and 2.5 kb. RT-PCR analysis showed that IGFBP-2 was expressed in all adult tissues studied, with the highest levels found in liver and skin. IGFBP-2 transcripts were detected in gonad during reproductive cycle of S. aurata. Highest levels of IGFBP-2 mRNA were found in bisexual young gonads, and the levels decreased with gonad development and relatively high levels of IGFBP-2 mRNA were found in the ovary during spawning. By contrast, testicular IGFBP-2 mRNA levels were very low. RT-PCR detected IGFBP-2 mRNA throughout development in unfertilized eggs, embryos, and larvae, with highest levels observed after day 3 post-hatching, suggesting that this mRNA is the product of both the maternal and embryonic genomes. High variability in steady-state levels of hepatic IGFBP-2 mRNA was noted in adult fish, resulting in a statistically insignificant response to growth hormone treatment. These results suggest that the structure of IGFBP-2 is conserved in sea bream and that IGFBP-2 mRNA is expressed during early development and in gonad during the reproductive cycle, suggesting that it may play a role in gilthead sea bream development and reproduction.
Funkenstein
B
B
National Institute of Oceanography, Israel Oceanographic and Limnological Research, Tel Shikmona, PO Box 8030, 31080, Haifa, Israel. bruria@ocean.org.il
Tsai
W
W
Maures
T
T
Duan
C
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D015870:Gene Expression / Q000187:drug effects; D018507:Gene Expression Regulation, Developmental; D013006:Growth Hormone / Q000494:pharmacology; D006801:Humans; D018971:Insulin-Like Growth Factor Binding Protein 2 / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D010473:Perciformes / Q000254:growth & development*; D010802:Phylogeny; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017385:Sequence Homology; D014018:Tissue Distribution
D018076:DNA, Complementary; D018971:Insulin-Like Growth Factor Binding Protein 2; D012333:RNA, Messenger; D013006:Growth Hormone