Single-molecule and super-resolved imaging deciphers membrane behavior of onco-immunogenic CCR5

Patrick Hunter; Alex L Payne-Dwyer; Michael Shaw; Nathalie Signoret; Mark C Leake

doi:10.1016/j.isci.2022.105675

Single-molecule and super-resolved imaging deciphers membrane behavior of onco-immunogenic CCR5

iScience. 2022 Nov 25;25(12):105675. doi: 10.1016/j.isci.2022.105675. eCollection 2022 Dec 22.

Authors

Patrick Hunter¹, Alex L Payne-Dwyer^{1

2}, Michael Shaw^{3

4}, Nathalie Signoret⁵, Mark C Leake^{1

2}

Affiliations

¹ Department of Biology, University of York, York YO10 5DD, UK.
² School of Physics, Engineering and Technology, University of York, York YO10 5DD, UK.
³ National Physical Laboratory, Hampton Road, Teddington, Middlesex TW11 0LW, UK.
⁴ Department of Computer Science, University College London, London WC1E 6EA, UK.
⁵ Hull York Medical School, University of York, York YO10 5DD, UK.

Abstract

The ability of tumors to establish a pro-tumorigenic microenvironment is an important point of investigation in the search for new therapeutics. Tumors form microenvironments in part by the "education" of immune cells attracted via chemotactic axes such as that of CCR5-CCL5. Further, CCR5 upregulation by cancer cells, coupled with its association with pro-tumorigenic features such as drug resistance and metastasis, has suggested CCR5 as a therapeutic target. However, with several conformational "pools" being reported, phenotypic investigations must be capable of unveiling conformational heterogeneity. Addressing this challenge, we performed super-resolution structured illumination microscopy (SIM) and single molecule partially TIRF-coupled HILO (PaTCH) microscopy of CCR5 in fixed cells. SIM data revealed a non-random spatial distribution of CCR5 assemblies, while Intensity-tracking of CCR5 assemblies from PaTCH images indicated dimeric sub-units independent of CCL5 perturbation. These biophysical methods can provide important insights into the structure and function of onco-immunogenic receptors and many other biomolecules.

Keywords: Biophysics; Cancer; Immunology.