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Article

Quantitative Super-Resolution Imaging for the Analysis of GPCR Oligomerization

1
London Centre for Nanotechnology, University College London, London WC1H 0AH, UK
2
Centre for Tumour Biology, Barts Cancer Institute, Queen Mary University of London, London EC1M 6BQ, UK
3
Centre for Endocrinology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London EC1M 6BQ, UK
4
Department of Chemistry, University College London, London WC1H 0AJ, UK
*
Author to whom correspondence should be addressed.
Academic Editors: Paolo Annibale and Marco Scarselli
Biomolecules 2021, 11(10), 1503; https://doi.org/10.3390/biom11101503
Received: 2 September 2021 / Revised: 6 October 2021 / Accepted: 9 October 2021 / Published: 12 October 2021
(This article belongs to the Special Issue Molecular Mechanisms of Compartmentalized GPCR Signaling)
G-protein coupled receptors (GPCRs) are known to form homo- and hetero- oligomers which are considered critical to modulate their function. However, studying the existence and functional implication of these complexes is not straightforward as controversial results are obtained depending on the method of analysis employed. Here, we use a quantitative single molecule super-resolution imaging technique named qPAINT to quantify complex formation within an example GPCR. qPAINT, based upon DNA-PAINT, takes advantage of the binding kinetics between fluorescently labelled DNA imager strands to complementary DNA docking strands coupled to protein targeting antibodies to quantify the protein copy number in nanoscale dimensions. We demonstrate qPAINT analysis via a novel pipeline to study the oligomerization of the purinergic receptor Y2 (P2Y2), a rhodopsin-like GPCR, highly expressed in the pancreatic cancer cell line AsPC-1, under control, agonistic and antagonistic conditions. Results reveal that whilst the density of P2Y2 receptors remained unchanged, antagonistic conditions displayed reduced percentage of oligomers, and smaller numbers of receptors in complexes. Yet, the oligomeric state of the receptors was not affected by agonist treatment, in line with previous reports. Understanding P2Y2 oligomerization under agonistic and antagonistic conditions will contribute to unravelling P2Y2 mechanistic action and therapeutic targeting. View Full-Text
Keywords: super-resolution; DNA-PAINT microscopy; qPAINT; G-protein coupled receptors; purinergic receptor Y2 (P2Y2); oligomerization super-resolution; DNA-PAINT microscopy; qPAINT; G-protein coupled receptors; purinergic receptor Y2 (P2Y2); oligomerization
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MDPI and ACS Style

Joseph, M.D.; Tomas Bort, E.; Grose, R.P.; McCormick, P.J.; Simoncelli, S. Quantitative Super-Resolution Imaging for the Analysis of GPCR Oligomerization. Biomolecules 2021, 11, 1503. https://doi.org/10.3390/biom11101503

AMA Style

Joseph MD, Tomas Bort E, Grose RP, McCormick PJ, Simoncelli S. Quantitative Super-Resolution Imaging for the Analysis of GPCR Oligomerization. Biomolecules. 2021; 11(10):1503. https://doi.org/10.3390/biom11101503

Chicago/Turabian Style

Joseph, Megan D., Elena Tomas Bort, Richard P. Grose, Peter J. McCormick, and Sabrina Simoncelli. 2021. "Quantitative Super-Resolution Imaging for the Analysis of GPCR Oligomerization" Biomolecules 11, no. 10: 1503. https://doi.org/10.3390/biom11101503

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