An international consortium of scientists, participated in by researchers from the University of Barcelona and the Bellvitge Biomedical Research Institute (IDIBELL), has developed the most extensive map to date on the interactions of G protein-coupled receptors (GPCR) in human beings. Through their results, published in Molecular Systems Biology, researchers have identified 987 unique interactions that occur among 686 proteins, 299 of which are integral membrane proteins. Furthermore, to demonstrate the biological relevance of the interactome, scientists validated novel interactions of the GPR37, serotonin 5‐HT4d and adenosine ADORA2A receptors through orthogonal analysis, using techniques such as coimmunosuppression and BRET.
To determine the interactome of the G protein-coupled receptors, researchers used the modified membrane yeast two-hybrid (MYTH) approach, a version of the classical double hybrid in yeast system. The application of this tool, which is not completely novel, as it has been used to identify different interactomes in yeasts or human cells, enables researchers to overcome certain limitations that other systems face such as the yeast two hybrid (YTH) system and tandem affinity purification combined with mass spectrometry (TAP/MS). In these two cases, cellular surroundings may be affected, altering the proteins and their interactions with other molecules, which is why scientists decided to use the first method to avoid modifications. Furthermore, according to a previous article published in Molecular Systems Biology in 2015, the MYTH method came with a lower price tag. In addition to that, it was simple and scalable, advantageous traits that made it ideal to analyze binary interactions.
“Thanks to the MYTH protocol, we have been able to study whole integral membrane proteins, such as the GPCR, in their membrane cellular surroundings. Among other results, researchers have defined the interactome of 48 different GPCRs with the MYTH technique,” says Francisco Ciruela, head of the Neuropharmacology and Pain Research Group of the University of Barcelona. According to scientists responsible for the study, later they conducted an “in-depth bioinformatics analysis to create and annotate an interactome for 48 selected full-length, clinically relevant human GPCRs in their ligand-unoccupied state, localized to their native plasma membrane.” Once this information was generated, the team carried out a systemic computational analysis to identify the best candidates to characterize their functional activity later on. Last, researchers studied the biological context of the interactome obtained, analyzing “its enrichment for pathways, diseases, molecular function, biological processes, domains, and drug targets.”
Source: Stagljar et al., Molecular Systems Biology 2017
Twenty-eight of the 48 G protein-coupled receptors are now therapeutic targets for a wide range of drugs, such as antihistaminics, Parkinson’s drugs or anti-psychotics. Overall, scientists estimate that the drugs that target these molecules stand for a 14 billion dollar market, thanks to the 27 million annual prescriptions written for them. This underscores the medical and economic importance of understanding in greater detail the interactions among these proteins. The most recent studies performed using proteomics techniques “do not make use of full‐length GPCRs in a natural cellular context”, according to the study’s authors, although the G protein-coupled receptors have indubitable biological relevance thanks to the role they play in cell signaling.
The new map developed on the interactions of GPCRs stands for another step towards determining in the future the origin of some of the most important diseases, such as schizophrenia, Parkinson’s, Alzheimer’s or epilepsy. By understanding how the various proteins connect in a network, scientists will research new therapeutic targets in the fight against this type of pathologies, as is shown by the fact that over half of the G protein-coupled receptors are already targets for a wide range of drugs. Thus, since Melanie L. Mayer and Philip Hieter published in 2000 the first interactome in a study published in Nature Biotechnology, understanding of these molecular interactions has only increased. This work enables scientists to clarify the networks that make up these proteins, so highly relevant in biology which, as they are immersed in the membranes of the cells participating in signal transduction, are difficult to analyze. The study, participated in by scientists of the UB and IDIBELL, will certainly “provide a valuable resource for the analysis of signaling pathways involving this druggable family of integral membrane proteins.”