Periodic Reporting for period 1 - GPCR Transcriptomics (Exploring the role of GPCR expression diversity in receptor physiology and drug response)
Periodo di rendicontazione: 2019-05-01 al 2021-04-30
G protein-coupled receptors (GPCRs) are a family of membrane proteins involved in the detection of extracellular signals such as light, odour, metabolites, neurotransmitters and hormones. In parallel with their importance as sensors of a range of extremely diverse signals that reach human cells and tissues, GPCRs are also the target of the majority of drugs available in the clinic. However, despite their importance in the detection of environmental cues, cell to cell communication and drug action, there are still several unresolved questions on receptor function that hamper our ability to fully understand their mechanisms of action. One of these questions is related to the fact that activation of the same receptor can lead to differential signalling responses depending on the cell, tissue or individual that is being studied. These discrepancies in receptor signalling can make it difficult to translate measurements in cells and model organisms to humans, to develop new GPCR drugs with improved therapeutic profiles, and to explain variability in receptor function across the population. The overall objective of this project was to apply data-driven computational analyses to determine how changes in receptors at the cell, tissue and individual level could lead to a diversification of GPCR signals.
To address this objective, we first used a set of integrative approaches combining data on human transcriptomics, genomics, proteomics, receptor sequence and structure, and molecular pharmacology to analyse how the tissue-specific expression of structurally and functionally diverse versions of the same receptor, in the form of GPCR protein isoforms, could contribute to differential signalling responses. Our results, which have been disseminated in conference presentations, through a high impact publication (Marti-Solano et al, Nature 2020), in the form of a dedicated press release (see link below) and via the deposition of our data in a web resource hosted by the widely used GPCR database in collaboration with researchers at the University of Copenhagen (gpcrdb.org/protein/isoforms) revealed that 38% of the GPCRs expressed in 30 different human tissues had two or more isoforms. Importantly, these isoforms displayed structural diversity in key receptor regions tasked with signal recognition (the N-terminal segment) and receptor signalling transmission and localisation (the C-terminal segment). Structural comparison between isoforms that had been functionally interrogated in the literature and previously uncharacterised isoforms also allowed proposing how variation in particular receptor regions could result in specific receptor signalling outcomes. Besides that, considering combinatorial expression of receptor isoforms across human tissues allowed determining which receptors could present a higher degree of functional diversity when exposed to a systemically-distributed signal or drug. Furthermore, this analysis also allowed detecting structurally diverse receptor isoforms with differential tissue distribution. This could open new opportunities for the development of next-generation isoform-specific GPCR drugs with an increased tissue selectivity that would be devoid of unwanted side effects in other unintended sites of action. In this line, external experimental collaborators at the universities of Michigan, Cambridge and Glasgow pharmacologically characterised the signalling properties of different receptor isoforms of pharmaceutical interest and validated our observations of signal diversification via the combinatorial expression of receptor isoforms in different cell systems. Importantly, during the timeline of the project, our data-driven methods also allowed us to analyse other sources of GPCR functional diversity related to the effects of rare variants found in the general population on receptor structure and signalling (Brouwers et al, Cell Rep 2021) and to changes in receptor abundance related to age and sex differences (manuscript in preparation).
In conclusion, our work revealed extensive expression and structural diversity in GPCRs that could help clarify previously unexplained differences in receptor signalling. Notably, the main results from this project, regarding signalling diversity resulting from receptor isoform expression, could have multiple implications in GPCR research and drug development. On the one hand, they point to the importance of ensuring that experimental models include the isoform expression profile of the cells and tissues that we intend to characterise or drug to facilitate the translational power of cellular and model organism readouts into insights on human physiology and therapeutic response. On the other, the observed isoform diversity could help clarify past tissue and cell-specific observations on drug action, and represent a new source of tissue and cell specificity via the targeting of drugs towards a given isoform of interest. Taken together, our recent publication should help guide new experimental efforts, both in academia and the pharmaceutical industry, to clarify the contribution of specific receptor isoforms to previously unexplained observations on receptor physiology and drug response, and could facilitate the development of more efficacious and safer GPCR drugs accounting for and exploiting isoform selectivity.