Asymmetric Recruitment of β-Arrestin1/2 by the Angiotensin II Type I and Prostaglandin F2α Receptor Dimer

Initially identified as monomers, G protein-coupled receptors (GPCRs) can also form functional homo- and heterodimers that act as distinct signaling hubs for cellular signal integration. We previously found that the angiotensin II (Ang II) type 1 receptor (AT1R) and the prostaglandin F2 alpha (PGF2 alpha) receptor (FP), both important in the control of smooth muscle contractility, form such a functional heterodimeric complex in HEK 293 and vascular smooth muscle cells. Here, we hypothesize that both Ang II- and PGF2 alpha-induced activation of the AT1R/FP dimer, or the parent receptors alone, differentially regulate signaling by distinct patterns of beta-arrestin recruitment. Using BRET-based biosensors, we assessed the recruitment kinetics of beta-arrestin1/2 to the AT1R/FP dimer, or the parent receptors alone, when stimulated by either Ang II or PGF2 alpha. Using cell lines with CRISPR/Cas9-mediated gene deletion, we also examined the role of G proteins in such recruitment. We observed that Ang II induced a rapid, robust, and sustained recruitment of beta-arrestin1/2 to AT1R and, to a lesser extent, the heterodimer, as expected, since AT1R is a strong recruiter of both beta-arrestin subtypes. However, PGF2 alpha did not induce such recruitment to FP alone, although it did when the AT1R is present as a heterodimer. beta-arrestins were likely recruited to the AT1R partner of the dimer. G alpha(q), G alpha(11), G alpha(12), and G alpha(13) were all involved to some extent in PGF2 alpha-induced beta-arrestin1/2 recruitment to the dimer as their combined absence abrogated the response, and their separate re-expression was sufficient to partially restore it. Taken together, our data sheds light on a newmechanismwhereby PGF2 alpha specifically recruits and signals through beta-arrestin but only in the context of the AT1R/FP dimer, suggesting that this may be a new allosteric signaling entity.