Molecular cloning and characterization of two Juvenile hormone esterase-like carboxylesterases cDNAs from the Morotoge shrimp, Pandalopsis japonica

Abstract

Methyl farnesoate (MF), a crustacean juvenile hormone (JH) analog, plays important roles in the regulation of a number of physiological processes such as molting, metamorphosis, reproductive maturation. Understanding its metabolic pathways is a key for various potential applications in crustacean aquaculture including artificial seed production and facilitation of growth and molt. Synthetic pathway of MF has been relatively well established but little has been known about its degradation and recycling pathways in crustacean. In insect juvenile hormone esterase (JHE) appeared to be responsible for JH metabolism. In order to expand our knowledge about MF metabolism, we isolated two cDNAs encoding JHE-like carboxylesterases (CXEs) from the hepatopancreas and ovary of Pandalopsis japonica by combination of the in-silico data mining from the expressed sequence tag (EST) database and traditional PCR-based cloning strategy. Full length of Pj-CXE1 (2084bp) and Pj-CXE2 (1985bp) cDNAs encoded a protein composed of 584 and 581 amino acids residues respectively. Structural analysis with other known carboxylesterases from various species revealed that the active sites and domain organizations of Pj-CXEs were well conserved suggesting that they may act on the similar substrate as shown in other carboxylesterases. Monophylic relationship among Pj-CXEs and other JHEs and acetylcholine esterases (AchEs) was not constructed by phylogenetic analysis suggesting that evolutional relationship may be more complicated than our thoughts. End-point RT-PCR results showed that Pj-CXE1 was predominantly expressed in the gonad whereas Pj-CXE2 was highly expressed in both the hepatopancreas and hindgut. Quantitative PCR results found that expression of Pj-CXE1 was upregulated in the gonads from the 7 days post eyestalk ablation (ESA) group. In contrast, no significant expressional change of Pj-CXE2 has been identified in neither hepatopancreas nor gonad. Our study provided two novel decapods JHE-like CXE cDNAs and expression of Pj-CXE1 is related with factors from X-organ/sinus gland complex. Further study will be needed to expand our knowledge in MF metabolism and its role in crustaceans molting and reproduction.