Identification and Characterization of an Insulin-Like Receptor Involved in Crustacean Reproduction

• Published in: Endocrinology (Philadelphia) 2016-02, Vol.157 (2), p.928-941
• Main Authors: Sharabi, O, Manor, R, Weil, S, Aflalo, E. D, Lezer, Y, Levy, T, Aizen, J, Ventura, T, Mather, P. B, Khalaila, I, Sagi, A
• Format: Article
• Language: English
• Subjects: Animals; Crustacea; Crustaceans; Differentiation; DNA, Complementary; Female; Gene expression; Gene Library; Gonadal Hormones - metabolism; Gonads; Gonads - metabolism; Hypertrophy; Insulin; Ligands; Macrobrachium; Macrobrachium rosenbergii; Maintenance; Male; Males; Palaemonidae - genetics; Prawns; Receptor Protein-Tyrosine Kinases - genetics; Receptor Protein-Tyrosine Kinases - metabolism; Receptor, Insulin - genetics; Receptor, Insulin - metabolism; Receptors; Reproduction - genetics; RNA Interference; RNA-mediated interference; Sex differentiation; Sex Differentiation - genetics; Sperm; Transcriptomics
• ISSN: 0013-7227; 1945-7170
• DOI: 10.1210/en.2015-1391

Sexual differentiation and maintenance of masculinity in crustaceans has been suggested as being regulated by a single androgenic gland (AG) insulin-like peptide (IAG). However, downstream elements involved in the signaling cascade remain unknown. Here we identified and characterized a gene encoding an insulin-like receptor in the prawn Macrobrachium rosenbergii (Mr-IR), the first such gene detected in a decapod crustacean. In mining for IRs and other insulin signaling-related genes, we constructed a comprehensive M. rosenbergii transcriptomic library from multiple sources. In parallel we sequenced the complete Mr-IR cDNA, confirmed in the wide transcriptomic library. Mr-IR expression was detected in most tissues in both males and females, including the AG and gonads. To study Mr-IR function, we performed long-term RNA interference (RNAi) silencing in young male prawns. Although having no effect on growth, Mr-IR silencing advanced the appearance of a male-specific secondary trait. The most noted effects of Mr-IR silencing were hypertrophy of the AG and the associated increased production of Mr-IAG, with an unusual abundance of immature sperm cells being seen in the distal sperm duct. A ligand blot assay using de novo recombinant Mr-IAG confirmed the existence of a ligand-receptor interaction. Whereas these results suggest a role for Mr-IR in the regulation of the AG, we did not see any sexual shift after silencing of Mr-IR, as occurred when the ligand-encoding Mr-IAG gene was silenced. This suggests that sexual differentiation in crustaceans involve more than a single Mr-IAG receptor, emphasizing the complexity of sexual differentiation and maintenance.