Multiplexed analysis of the secretin-like GPCR-RAMP interactome
Open Access
- 6 September 2019
- journal article
- research article
- Published by American Association for the Advancement of Science (AAAS) in Science Advances
- Vol. 5 (9), eaaw2778
- https://doi.org/10.1126/sciadv.aaw2778
Abstract
Receptor activity–modifying proteins (RAMPs) have been shown to modulate the functions of several G protein–coupled receptors (GPCRs), but potential direct interactions among the three known RAMPs and hundreds of GPCRs have never been investigated. Focusing mainly on the secretin-like family of GPCRs, we engineered epitope-tagged GPCRs and RAMPs, and developed a multiplexed suspension bead array (SBA) immunoassay to detect GPCR-RAMP complexes from detergent-solubilized lysates. Using 64 antibodies raised against the native proteins and 4 antibodies targeting the epitope tags, we mapped the interactions among 23 GPCRs and 3 RAMPs. We validated nearly all previously reported secretin-like GPCR-RAMP interactions, and also found previously unidentified RAMP interactions with additional secretin-like GPCRs, chemokine receptors, and orphan receptors. The results provide a complete interactome of secretin-like GPCRs with RAMPs. The SBA strategy will be useful to search for additional GPCR-RAMP complexes and other interacting membrane protein pairs in cell lines and tissues.Funding Information
- Knut och Alice Wallenbergs Stiftelse
- Science for Life Laboratory
- Erling-Persson Family Foundation
- Nicholson Short-Term Exchange
- Albert Cass Fellowship
- Alexander Mauro Fellowship
- Danica Foundation
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