Neuropeptides are tiny messenger molecules used by neurons to control physiological functions and behaviour. Some of these neuropeptides are hormones that play a key role in the control of reproduction. In vertebrates,these neuropeptides include gonadotropin-releasing hormones (GnRHs), which regulate fertility through follicular growth and ovulation. The GnHRs bind to special receptors, which transmit the hormone's signal from the surface of the target cell to the interior, where the cell responds according to the message carried by the hormone. Genes encoding GnRH-like peptides and their receptors have also been identified in various invertebrates, including nematodes and insects, but their role in invertebrate reproduction was unclear. In a recent Proceedings of the National Academy of Sciences of the USA paper an international team of scientist led by Liliane Schoofs provides the first direct evidence that GnRH-mediated signalling has a role in invertebrate reproduction, and shows that this pathway is evolutionarily conserved from worms to humans.
Several years ago molecular biologists identified the first ligand of the insect GnRH receptor, the adipokinetic hormone (AKH), which is an invertebrate neuropeptide related to vertebrate GnRHs and is known to control energy metabolism. Although a role for AKH in invertebrate reproduction has been suggested, experimental proof was missing. It needed the help of a little worm, the nematode Caenorhabditis elegans, to confirm this assumption. In C. elegans, a GnRH receptor homologue had been reported previously, but it was thought to be an orphan receptor because no one had identified an AKH/GnRH-like neuropeptide that could bind to the receptor in the worm's genome.
To identify the hormone that binds the worm's GnRH receptor, Schoofs' team first tested whether the receptor is capable of binding AKH from another invertebrate, the fruit fly Drosophila. They cloned the worm receptor and expressed it in human cells. To demonstrate that the fly neuropeptide binds and activates the worm receptor, they measured the calcium concentration in the human cell's cytoplasm, as the calcium level is known to increase when the receptor is activated by binding its hormone. The Drosophila AKH peptide triggered an increase of the calcium concentration suggesting that an AKH/GnRH-related peptide is the natural ligand of the worm GnRH receptor.
Motivated by this finding, the scientists tried to identify an AKH/GnRH gene in the C. elegans genome. By improving their search scheme, they finally discovered a gene encoding a neuropeptide that was strikingly similar to arthropod AKHs and vertebrate GnRHs. The next step was to figure out whether the newly discovered AKH/GnRH-related peptide also activates the worm's GnRH receptor. Synthesizing the peptide, the team tested it at different concentrations to see whether it binds to the worm receptor and triggers the calcium response, and found that it did, even at low concentrations.
But the scientists still did not know the biological function of the newly discovered neuropeptide. To get some clues to its putative role in reproduction, the team knocked out expression of the genes encoding the AKH/GnRH-related peptide and its receptor, and analysed the worm's ability to produce offspring. The worm's fertility was affected. The scientists observed a delay in egg laying and a decrease in the number of progeny.
Liliane Schoofs' team has provided the first clear evidence that an AKH/GnRH-related peptide and its receptor have a role in the control of reproduction in invertebrates. It seems that the fundamental concepts in the regulation of reproduction are highly conserved throughout the animal kingdom. Obviously, the underlying signalling machinery evolved very early in metazoan evolution.