(819.8) Membrane Phosphoinositides Stabilize GPCR-arrestin Complexes and Provide Temporal Control of Complex Assembly and Dynamics

Poster Board Number: A432

John Janetzko (Stanford University), Ryoji Kise (Tohoku University), Benjamin Barsi-Rhyne (University of California San Francisco), Dirk Siepe (Stanford University), Franziska Heydenreich (Stanford University), Matthieu Masureel (Stanford University), Kouki Kawakami (Tohoku University), K. Garcia (Stanford University), Mark von Zastrow (University of California San Francisco), Asuka Inoue (Tohoku University), Brian Kobilka (Stanford University)

Binding of arrestin to phosphorylated G protein-coupled receptors (GPCRs) is crucial for gating signaling. Once internalized some GPCRs remain stably associated with arrestin, while others interact transiently; this difference affects signaling and recycling behaviors of these GPCRs. Using cell-based and in vitro biophysical assays we examined the role of membrane phosphoinositides (PIPs) in arrestin recruitment and GPCR-arrestin complex dynamics. We find that GPCRs broadly stratify into two groups, one which requires PIP-binding for arrestin recruitment and one that does not. Plasma membrane PIPs potentiate an active conformation of arrestin and stabilize GPCR-arrestin complexes by promoting a core-engaged state of the complex. As allosteric modulators of GPCR-arrestin complex dynamics, membrane PIPs allow for additional conformational diversity beyond that imposed by GPCR phosphorylation alone. The dependance on membrane PIPs provides a mechanism for arrestin release from transiently associated GPCRs, allowing their rapid recycling, while explaining how stably associated GPCRs are able to engage G proteins at endosomes.

This work was supported in part by National Institutes of Health grants R01NS028471 (B.K.K.), R01 AI125320 (K.C.G), R01DA010711 and R01DA012864 (M.vZ.). Additional support to both K.C.G and B.K.K. is provided by the Mathers Foundation. B.K.K. is a Chan-Zuckerberg Biohub Investigator. J.J. is a Damon Runyon Fellow supported by the Damon Runyon Cancer Research Foundation (DRG-2318-18). B.B.R. is a recipient of an American Heart Association Predoctoral Fellowship (19PRE34380570). F.M. H. is a recipient of a Marie Skłodowska-Curie Individual Fellowship from the European Unionamp;rsquo;s Horizon 2020 research and innovation programme (grant agreement No. 844622) and an American Heart Association postdoctoral fellowship (19POST34380839). M.M. was supported by an American Heart Association postdoctoral fellowship (17POST33410958). A.I. was funded by the PRIME 19gm5910013, the LEAP 20gm0010004 and the BINDS JP20am0101095 from the Japan Agency for Medical Research and Development (AMED); KAKENHI 17K08264 and 21H04791 from by the Japan Society for the Promotion of Science (JSPS); JST Moonshot Research and Development Program JPMJMS2023 from Japan Science and Technology Agency (JST); Daiichi Sankyo Foundation of Life Science; Takeda Science Foundation; Ono Medical Research Foundation; The Uehara Memorial Foundation.