LUXendins reveal endogenous glucagon-like peptide-1 receptor distribution and dynamics
Ast J., Arvaniti A., Fine NHF., Nasteska D., Ashford FB., Stamataki Z., Koszegi Z., Bacon A., Trapp S., Jones BJ., Hastoy B., Tomas A., Reissaus CA., Linnemann AK., D’Este E., Calebiro D., Johnsson K., Podewin T., Broichhagen J., Hodson DJ.
<jats:title>ABSTRACT</jats:title><jats:p>The glucagon-like peptide-1 receptor (GLP1R) is a class B G protein-coupled receptor (GPCR) involved in metabolism. Presently, its visualization is limited to genetic manipulation, antibody detection or the use of probes that stimulate receptor activation. Herein, we present <jats:bold>LUXendin645</jats:bold>, a far-red fluorescent GLP1R antagonistic peptide label. <jats:bold>LUXendin645</jats:bold> produces intense and specific membrane labeling throughout live and fixed tissue. GLP1R signaling can additionally be evoked when the receptor is allosterically modulated in the presence of <jats:bold>LUXendin645</jats:bold>. Using <jats:bold>LUXendin645</jats:bold> and STED-compatible <jats:bold>LUXendin651</jats:bold> we describe islet GLP1R expression patterns, reveal higher-order GLP1R organization including the existence of membrane nanodomains, and track single receptor subpopulations. We furthermore show that different fluorophores can confer agonistic behavior on the <jats:bold>LUXendin</jats:bold> backbone, with implications for the design of stabilized incretin-mimetics. Thus, our labeling probes possess divergent activation modes, allow visualization of endogenous GLP1R, and provide new insight into class B GPCR distribution and dynamics.</jats:p>