Investigating a small class of five receptors in Drosophila two years ago, Paul Taghert's group at Washington University School of Medicine discovered that one of these is a receptor for diuretic hormone 44, which regulates osmotic balance. Suspecting that some of the other four receptors in this class may play similar physiological roles, they embarked on an international collaboration with groups led by Julian Dow and David Schooley to search for the ligand for another receptor in this class, CG17415(p. 1239). Taghert explains that the interaction between a receptor protein and its peptide ligand is very specific, like a lock and key, causing a change in the receptor protein that triggers a series of intracellular signals. In some cases accessory proteins, which assist in receptor–ligand binding, need to be present for receptors to function properly. Drosophila's CG17415 receptor is closely related to a mammalian receptor that needs accessory proteins to work, so the team reasoned that this might also be the case for the fruit fly receptor.
The group set out to identify which unique ligand pairs up with the CG17415 receptor, and to see whether it needs accessory proteins. They transfected the CG17415 receptor gene into mammalian tissue cultures so that these cells now expressed the gene and contained the receptor in their cell membrane, acting like an antenna for the right ligand. When a ligand binds to its receptor, the concentration of a messenger molecule (in this case cyclic AMP) increases. If the team saw an accumulation of this messenger after adding a potential ligand to the cell cultures, they would know that they had found the right ligand. They exposed the cells to 23 potential peptide ligands, but did not see an accumulation of cyclic AMP in response to any of these until they added mammalian accessory proteins to the cells. With the accessory proteins assisting the process, the team saw a clear response to just one of the 23 peptides: diuretic hormone 31. Clearly, CG17415 is a diuretic hormone 31 receptor, and just as they had suspected, it needs accessory proteins to work properly.
But the team still didn't know where this receptor is expressed in the fruit fly. If CG17415 is a diuretic hormone receptor, the team reasoned that they should find the receptor in the fruit fly's `kidney', the Malpighian tubules, which regulate fluid balance. Sure enough, when the team used antibodies for the CG17415 receptor to stain and localise the receptor protein, they saw that the receptor is expressed in the tubules.
The immunostaining also revealed something unexpected. The team were surprised to find that a small cluster of 30 neurons in Drosophila's brain express both the diuretic hormone 31 receptor and the diuretic hormone 44 receptor that the team discovered two years ago, which suggests that these two hormones have convergent signalling pathways. Taghert explains that the diuretic hormone 31 receptor is related to the mammalian calcitonin gene-related peptide (CGRP) receptor, while the diuretic hormone 44 receptor is related to the mammalian corticotrophin-releasing factor (CRF) receptor.`Co-expression of CGRP and CRF has also been suggested in mammals,' says Taghert. He concludes, `the convergence of these two receptors in fruit fly brains and mammalian brain circuitry suggests striking evolutionary parallels between insect and mammalian hormone signalling pathways.'
