Cloning and analysis of the lamprey GnRH-III cDNA from eight species of lamprey representing the three families of Petromyzoniformes

Abstract

The lamprey, which are divided into three families, including the Petromyzonidae, Geotriidae, and Mordaciidae, have been shown to regulate the reproductive axis through a functional hypothalamic–pituitary–gonadal axis. To date, two forms of gonadotropin-releasing hormone (GnRH) have been identified in the sea lamprey ( Petromyzon marinus), lamprey GnRH-I (decapeptide and cDNA) and lamprey GnRH-III (decapeptide), both of which have been shown to be expressed in the preoptic-anterior hypothalamic region and both forms have been demonstrated to regulate reproductive function (i.e. steroidogenesis and gametogenesis). The objective of this study was to isolate the cDNA encoding the prepro-lamprey GnRH-III from eight species of lamprey using a PCR based subcloning procedure. A degenerate primer designed to the lamprey GnRH-III decapeptide was used to amplify the 3′ end of each transcript, while gene specific primers were used to amplify the 5′ ends. Phylogenetic analysis using the prepro-lamprey GnRH-III amino acid sequences was performed, in which the lamprey GnRH-III sequences divided into three groups, supporting the current view of the lamprey lineage at the family level. Finally, a phylogenetic analysis of these newly identified deduced amino acid sequences together with 64 previously described GnRH sequences suggests that the lamprey GnRHs are unique, as they group together separately from the three previously described paralogous lineages of the GnRH family.