Multiplexed analysis of the secretin-like GPCR-RAMP interactome

Lorenzen, EmilyRockefeller Univ, Lab Chem Biol & Signal Transduct, 1230 York Ave, New York, NY 10065 USA. (författare)

QC 20191115
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.

Dodig-Crnkovic, TeaKTH,Affinitets-proteomik,Science for Life Laboratory, SciLifeLab(Swepub:kth)u1wrth1x (författare)
Kotliar, Ilana B.Rockefeller Univ, Lab Chem Biol & Signal Transduct, 1230 York Ave, New York, NY 10065 USA.;Tri Inst PhD Program Chem Biol, New York, NY 10065 USA. (författare)
Pin, ElisaKTH,Affinitets-proteomik,Science for Life Laboratory, SciLifeLab(Swepub:kth)u1pmjyje (författare)
Ceraudo, EmilieRockefeller Univ, Lab Chem Biol & Signal Transduct, 1230 York Ave, New York, NY 10065 USA. (författare)
Vaughan, Roger D.Rockefeller Univ, Ctr Clin & Translat Sci, 1230 York Ave, New York, NY 10065 USA. (författare)
Uhlén, MathiasKTH,Science for Life Laboratory, SciLifeLab,Systembiologi(Swepub:kth)u1dulvmw (författare)
Huber, ThomasRockefeller Univ, Lab Chem Biol & Signal Transduct, 1230 York Ave, New York, NY 10065 USA. (författare)
Schwenk, Jochen M.KTH,Science for Life Laboratory, SciLifeLab,Affinitets-proteomik(Swepub:kth)u1h7wtme (författare)
Sakmar, Thomas P.Karolinska Institutet (författare)
Rockefeller Univ, Lab Chem Biol & Signal Transduct, 1230 York Ave, New York, NY 10065 USA.Affinitets-proteomik (creator_code:org_t)

Av författaren/redakt...: Lorenzen, Emily; Dodig-Crnkovic, ...; Kotliar, Ilana B ...; Pin, Elisa; Ceraudo, Emilie; Vaughan, Roger D ...; visa fler...; Uhlén, Mathias; Huber, Thomas; Schwenk, Jochen ...; Sakmar, Thomas P ...; visa färre...

Om ämnet

MEDICIN OCH HÄLSOVETENSKAP: MEDICIN OCH HÄLS ...; och Medicinska och f ...; och Cell och molekyl ...

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