A study on the functional interaction between the GH/PRL family of polypeptides with their receptors in zebrafish: Evidence against GHR1 being the receptor for somatolactin.

The growth hormone (GH)/prolactin (PRL) family of polypeptide hormones plays important roles in many aspects of vertebrate physiology. In fish, there is an additional member in this family called somatolactin (SL). Specifically, zebrafish contains five ligands (GH, SLα, SLß, PRL1 and PRL2) and four cognate receptors including two GH receptors (GHR1 and GHR2) and two PRL receptors (PRLR1 and PRLR2). There is much controversy regarding whether one of the two GHRs in teleosts is in fact the receptor of SL. A multitude of different assay methods were employed to study the functional interaction among these ligands and their receptors in zebrafish. These include assessment of the binding between the ligands and the extracellular domains of the receptors using His-tag pulldown assays, activation of receptor-evoked promoter activities by treatment of the receptor-transfected cells with the recombinant hormones, and phosphorylation of post-receptor signaling factors by treatment of receptor-transfected cells with the recombinant hormones. The results showed that the zebrafish GH can only interact with the GHRs and the zebrafish PRLs can only interact with the PRLRs. The zebrafish SLs, found to be biologically active in another assay, were found to be ineffective in interacting with the zebrafish GHRs and PRLRs. Our data argue against the hypothesis that GHR1 is the SL receptor.

Discovery of a novel prolactin in non-mammalian vertebrates: evolutionary perspectives and its involvement in teleost retina development.

BACKGROUND: The three pituitary hormones, viz. prolactin (PRL), growth hormone (GH) and somatolactin (SL), together with the mammalian placental lactogen (PL), constitute a gene family of hormones with similar gene structure and encoded protein sequences. These hormones are believed to have evolved from a common ancestral gene through several rounds of gene duplication and subsequent divergence. PRINCIPAL FINDINGS: In this study, we have identified a new PRL-like gene in non-mammalian vertebrates through bioinformatics and molecular cloning means. Phylogenetic analyses showed that this novel protein is homologous to the previously identified PRL. A receptor transactivation assay further showed that this novel protein could bind to PRL receptor to trigger the downstream post-receptor event, indicating that it is biologically active. In view of its close phylogenetic relationship with PRL and also its ability to activate PRL receptor, we name it as PRL2 and the previously identified PRL as PRL1. All the newly discovered PRL2 sequences possess three conserved disulfide linkages with the exception of the shark PRL2 which has only two. In sharp contrast to the classical PRL1 which is predominantly expressed in the pituitary, PRL2 was found to be mainly expressed in the eye and brain of the zebrafish but not in the pituitary. A largely reduced inner nuclear layer of the retina was observed after morpholino knockdown of zebrafish PRL2, indicating its role on retina development in teleost. SIGNIFICANCE: The discovery of this novel PRL has revitalized our understanding on the evolution of the GH/PRL/SL/PL gene family. Its unique expression and functions in the zebrafish eye also provide a new avenue of research on the neuroendocrine control of retina development in vertebrates.