Molecular identification of a new androgenic gland-specific insulin-like gene from the mud crab, Scylla paramamosain

• A new insulin-like gene Sp-IAG was identified from Scylla paramamosain.
• The complete cDNA and genomic DNA with a 5′ flanking region sequences were revealed.
• Sp-IAG mRNA and protein dominantly expressed in the male reproductive organ
• The Sp-IAG mRNA expression was up-regulated at mating.
• Eyestalk ablation can induce high expression of Sp-IAG mRNA.

It is well known that the androgenic gland is a specific endocrine organ of the male crustacean, controlling its primary and secondary sexual characteristics. The androgenic gland hormone is a key peptide hormone involved in sexual differentiation in crustaceans. In this study, a full length cDNA encoding insulin-like gland hormone from Scylla paramamosain, named Sp-IAG, was identified. The predicted protein had similar molecular organization to other insulin-like androgenic gland factors reported in the Decapoda, encoding a signal peptide, B chain, C peptide and A chain. There were two disulfide bridges between the B chain and A chain, and one disulfide in the B chain. In particular, a full genomic DNA sequence of Sp-IAG with a 5′ flanking region of 2988 bp was revealed from which many potential regulatory binding sites were first identified in S. paramamosain. The Sp-IAG gene was highly expressed in the androgenic gland and also lowly expressed in the seminal vesicle and ejaculatory duct, which was not reported in previous studies. It is noteworthy that Sp-IAG was highly expressed both in the androgenic gland and in the seminal vesicle using western blot analysis, suggesting that Sp-IAG might play a major role in these two organs. In addition, it was quantitatively detected that Sp-IAG mRNA expression was up-regulated after mating, which suggested that this protein might have an activity related to mating behavior. Eyestalk ablation can induce a high level of Sp-IAG expression, demonstrating that the Sp-IAG of S. paramamosain was negatively controlled by the X-organ–sinus gland in the eyestalk. These findings provided the basis for elucidating the mechanism of sex differentiation in crustaceans.