Abstract
The sea cucumber Holothuria scabra is an economically valuable marine species which is distributed throughout the Asia-Pacific region. With the natural population declining due to over fishing, aquaculture of this species is deemed necessary. Hence, it is essential to understand the mechanisms regulating the reproduction in order to increase their populations. Sex steroids, including estrogens, androgens and progestogens, play an important role in reproduction in most vertebrates and several invertebrates. It has been proposed that sea cucumbers have the same sex steroids as vertebrates but the steroidogenic pathway in the sea cucumbers is still unclear. In this study, we demonstrated by using liquid chromatography-tandem mass spectrometry (LC-MS/MS) that sex steroids (estradiol, progesterone, and testosterone) were present in H. scabra neural and gonadal tissues. In silico searches of available sea cucumber transcriptome data identified 26 steroidogenesis-related genes. Comparative analysis of encoded proteins for the steroidogenic acute regulatory protein (HscStAR), CYP P450 10, 17 and 3A (HscCYP10, HscCYP17, HscCYP3A) and hydroxysteroid dehydrogenases (Hsc3β-HSD, Hsc17β-HSD) with other species was performed to confirm their evolutionary conservation. Gene expression analyses revealed widespread tissue expression. Real-time PCR analysis revealed that HscStAR, HscCYP10, Hsc3β-HSD, and Hsc17β-HSD gene expressions were similar to those in ovaries and testes, which increased during the gonad maturation. HscCYP17 mRNA was increased during ovarian development and its expression declined at late stages in females but continued high level in males. The expression of the HscCYP3A was high at the early stages of ovarian development, but not at other later stages in ovaries, however it remained low in testes. Moreover, a role for steroids in reproduction was confirmed following the effect of sex steroids on vitellogenin (Vtg) expression in ovary explant culture, showing upregulation of Vtg level. Collectively, this study has confirmed the existence of steroids in an echinoderm, as well as characterizing key genes associated with the steroidogenic pathway. We propose that sex steroids might also be associated with the reproduction of H. scabra, and the identification of biosynthetic genes enables future functional studies to be performed.