Gs is dispensable for (3-arrestin coupling but dictates GRK selectivity and is predominant for gene expression regulation by (32-adrenergic receptor

beta-arrestins play a key role in G protein-coupled receptor (GPCR) internalization, trafficking, and signaling. Whether beta- arrestins act independently of G protein-mediated signaling has not been fully elucidated. Studies using genome-editing approaches revealed that whereas G proteins are essential for mitogen-activated protein kinase activation by GPCRs., beta- arrestins play a more prominent role in signal compartmentalization. However, in the absence of G proteins, GPCRs may not activate beta-arrestins, thereby limiting the ability to distinguish G protein from beta-arrestin-mediated signaling events. We used beta 2-adrenergic receptor (beta 2AR) and its beta 2AR-C tail mutant expressed in human embryonic kidney 293 cells wildtype or CRISPR-Cas9 gene edited for G alpha s, beta-arrestin1/2, or GPCR kinases 2/3/5/6 in combination with arrestin conformational sensors to elucidate the interplay between G alpha s and beta-arrestins in controlling gene expression. We found that G alpha s is not required for beta 2AR and beta-arrestin conformational changes, beta-arrestin recruitment, and receptor internalization, but that G alpha s dictates the GPCR kinase isoforms involved in beta-arrestin recruitment. By RNA-Seq analysis, we found that protein kinase A and mitogenactivated protein kinase gene signatures were activated by stimulation of beta 2AR in wildtype and beta-arrestin1/2-KO cells but absent in G alpha s-KO cells. These results were validated by re expressing G alpha s in the corresponding KO cells and silencing beta- arrestins in wildtype cells. These findings were extended to cellular systems expressing endogenous levels of beta 2AR. Overall, our results support that Gs is essential for beta 2AR-promoted protein kinase A and mitogen-activated protein kinase gene expression signatures, whereas beta-arrestins initiate signaling events modulating G alpha s-driven nuclear transcriptional activity.