PEGylated isoprenaline reveals distinct functions of cardiac β-adrenergic receptors located in the T-tubule vs . outer surface membrane

Marion Barthé (1) , Flora Lefebvre (2) , Emilie Langlois (2) , Florence Lefebvre (1) , Patrick Lechêne (1) , Xavier Iturrioz (3) , Tâp Ha-Duong (4) , Laurence Moine (2) , Nicolas Tsapis , Rodolphe Fischmeister (1)

1 CARPAT (UMRS1180) - Signalisation et physiopathologie cardiovasculaire
2 IGPS - Institut Galien Paris-Saclay
3 CIRB - Centre interdisciplinaire de recherche en biologie
4 BioCIS - Biomolécules : Conception, Isolement, Synthèse

Membrane proteins are present in both cardiac T-tubule (TTM) and outer surface membrane (OSM), although at a different density. Classical pharmacology does not allow to explore the function of a membrane protein separately in OSM vs. TTM. Here, we developed a technology based on size exclusion to explore the function of β-adrenergic receptors (β-ARs) located in the OSM. We synthetized a PEG-Iso molecule by covalent linking between isoprenaline (Iso) and a 5000 Da PolyEthylene-Glycol (PEG). Using confocal microscopy, we show that PEGylation constrains molecules outside the T-tubule network. PEG-Iso produced similar effects as Iso on I ca,L , sarcomere shortening and Ca 2+ transients. However, PEG-Iso increased [cAMP] i with a lower efficacy than Iso, produced a much lower stimulation of nuclear PKA activity than Iso but a larger stimulation of cytosolic PKA at equivalent levels of [cAMP] i . Our results show that activation of OSM β-ARs is sufficient to activate cytosolic PKA and excitation-contraction coupling, but insufficient to activate nuclear PKA or nuclear protein phosphorylation for which additional activation of TTM β-ARs is needed.

Domaines: Cardiologie et système cardiovasculaire

Marion Barthé, Flora Lefebvre, Emilie Langlois, Florence Lefebvre, Patrick Lechêne, et al.. PEGylated isoprenaline reveals distinct functions of cardiac β-adrenergic receptors located in the T-tubule vs . outer surface membrane. 2022. ⟨hal-03793970⟩