Engineered hyperphosphorylation of the b2- Adrenoceptor prolongs arrestin-3 binding and induces arrestin internalization

Diana Zindel, Adrian J. Butcher, Suleiman Al-Sabah, Peter Lanzerstorfer, Julian Weghuber, Andrew B. Tobin, Moritz Bünemann, Cornelius Krasel

Publikation: Beitrag in FachzeitschriftArtikelBegutachtung

23 Zitate (Scopus)

Abstract

G protein-coupled receptor phosphorylation plays a major role in receptor desensitization and arrestin binding. It is, however, unclear how distinct receptor phosphorylation patterns may influence arrestin binding and subsequent trafficking. Here we engineer phosphorylation sites into the C- Terminal tail of the β2- Adrenoceptor (β2AR) and demonstrate that this mutant, termed β2ARSSS, showed increased isoprenaline-stimulated phosphorylation and differences in arrestin-3 affinity and trafficking. By measuring arrestin-3 recruitment and the stability of arrestin-3 receptor complexes in real time using fluorescence resonance energy transfer and fluorescence recovery after photobleaching, we demonstrate that arrestin-3 dissociated quickly and almost completely from the β2AR, whereas the interaction with β2ARSSS was 2- To 4-fold prolonged. In contrast, arrestin-3 interaction with a β2- Adrenoceptor fused to the carboxyl- Terminal tail of the vasopressin type 2 receptor was nearly irreversible. Further analysis of arrestin-3 localization revealed that by engineering phosphorylation sites into the b2- Adrenoceptor the receptor showed prolonged interaction with arrestin-3 and colocalization with arrestin in endosomes after internalization. This is in contrast to the wild- Type receptor that interacts transiently with arrestin-3 at the plasma membrane. Furthermore, β2ARSSS internalized more efficiently than the wild- Type receptor, whereas recycling was very similar for both receptors. Thus, we show how the interaction between arrestins and receptors can be increased with minimal receptor modification and that relatively modest increases in receptor- Arrestin affinity are sufficient to alter arrestin trafficking.

OriginalspracheEnglisch
Seiten (von - bis)349-362
Seitenumfang14
FachzeitschriftMolecular Pharmacology
Jahrgang87
Ausgabenummer2
DOIs
PublikationsstatusVeröffentlicht - Feb. 2015

Fingerprint

Untersuchen Sie die Forschungsthemen von „Engineered hyperphosphorylation of the b2- Adrenoceptor prolongs arrestin-3 binding and induces arrestin internalization“. Zusammen bilden sie einen einzigartigen Fingerprint.

Zitieren