Single-molecule FRET imaging of GPCR dimers in living cells

Wesley B. Asher, Peter Geggier, Michael D. Holsey, Grant T. Gilmore, Avik K. Pati, Jozsef Meszaros, Daniel S. Terry, Signe Mathiasen, Megan J. Kaliszewski, Mitchell D. McCauley, Alekhya Govindaraju, Zhou Zhou, Kaleeckal G. Harikumar, Khuloud Jaqaman, Laurence J Miller, Adam W. Smith, Scott C. Blanchard, Jonathan A. Javitch*

*Corresponding author af dette arbejde

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

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Abstract

Class C G protein-coupled receptors (GPCRs) are known to form stable homodimers or heterodimers critical for function, but the oligomeric status of class A and B receptors, which constitute >90% of all GPCRs, remains hotly debated. Single-molecule fluorescence resonance energy transfer (smFRET) is a powerful approach with the potential to reveal valuable insights into GPCR organization but has rarely been used in living cells to study protein systems. Here, we report generally applicable methods for using smFRET to detect and track transmembrane proteins diffusing within the plasma membrane of mammalian cells. We leverage this in-cell smFRET approach to show agonist-induced structural dynamics within individual metabotropic glutamate receptor dimers. We apply these methods to representative class A, B and C receptors, finding evidence for receptor monomers, density-dependent dimers and constitutive dimers, respectively.

OriginalsprogEngelsk
TidsskriftNature Methods
Vol/bind18
Udgave nummer4
Sider (fra-til)397-405
Antal sider9
ISSN1548-7091
DOI
StatusUdgivet - 2021
Udgivet eksterntJa

Bibliografisk note

Publisher Copyright:
© 2021, The Author(s), under exclusive licence to Springer Nature America, Inc.

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