G protein alpha subunits and beta gamma dimers are covalently modified by lipids. The emerging picture is one in which attached lipids provide more than just a nonspecific "glue" for sticking G proteins to membranes. We are only beginning to understand how different lipid modifications of different G protein subunits affect specific protein-protein interactions and localization to specific cellular sites. In addition, regulation of these modifications, particularly palmitoylation, can provide new ways to regulate signals transmitted by G proteins.
؊ . This revealed the specific ability of bovine brain G␣ q/11 to bind to both GRK2 and GRK3 in an AlF 4 ؊ -dependent manner. In contrast, G␣ s , G␣ i , and G␣ 12/13 did not bind to GRK2 or GRK3 despite their presence in the extract. Additional studies revealed that bovine brain G␣ q/11 could also bind to an N-terminal construct of GRK2, while no binding of G␣ q/11 , G␣ s , G␣ i , or G␣ 12/13 to comparable constructs of GRK5 or GRK6 was observed. Experiments using purified G␣ q revealed significant binding of both G␣ q GDP/AlF 4 ؊ and G␣ q (GTP␥S), but not G␣ q (GDP), to GRK2. Activation-dependent binding was also observed in both COS-1 and HEK293 cells as GRK2 significantly co-immunoprecipitated constitutively active G␣ q (R183C) but not wild type G␣ q . In vitro analysis revealed that GRK2 possesses weak GAP activity toward G␣ q that is dependent on the presence of a G proteincoupled receptor. However, GRK2 effectively inhibited G␣ q -mediated activation of phospholipase C- both in vitro and in cells, possibly through sequestration of activated G␣ q . These data suggest that a subfamily of the GRKs may be bifunctional regulators of G protein-coupled receptor signaling operating directly on both receptors and G proteins.
G protein-coupled receptors (GPCRs) are increasingly recognized to operate from intracellular membranes as well as the plasma membrane. The β2-adrenergic GPCR can activate Gs-linkedcyclic AMP (cAMP) signaling from endosomes. We show here that the homologous human β1-adrenergic receptor initiates an internal Gs-cAMP signal from the Golgi apparatus. By developing a chemical method to acutely squelch G protein coupling at defined membrane locations, we demonstrate that Golgi activation contributes significantly to the overall cellular cAMP response. Golgi signalling utilizes a pre-existing receptor pool rather than receptors delivered from the cell surface, requiring separate access of extracellular ligands. Epinephrine, a hydrophilic endogenous ligand, accesses the Golgi-localized receptor pool by facilitated transport requiring the organic cation transporter 3 (OCT3) whereas drugs can access the Golgi pool by passive diffusion according to hydrophobicity. We demonstrate marked differences among both agonist and antagonist drugs in Golgi-localized receptor access, and show that β-blocker drugs presently used in the clinic differ markedly in ability to antagonize the Golgi signal. We propose ’location bias’ as a new principle for achieving functional selectivity of GPCR-directed drug action.
To be activated by cell surface G protein-coupled receptors, heterotrimeric G proteins must localize at the cytoplasmic surface of plasma membranes. Moreover, some G protein subunits are able to traffic reversibly from the plasma membrane to intracellular locations upon activation. This review will highlight new insights into how nascent G protein subunits are assembled and how they arrive at plasma membranes. In addition, recent reports have increased our knowledge of activation-induced trafficking of G proteins. Understanding G protein assembly and trafficking will lead to a greater understanding of novel ways that cells regulate G protein signaling.
Peripheral membrane proteins utilize a variety of mechanisms to attach tightly, and often reversibly, to cellular membranes. The covalent lipid modifications, myristoylation and palmitoylation, are critical for plasma membrane localization of heterotrimeric G protein alpha subunits. For alpha(s) and alpha(q), two subunits that are palmitoylated but not myristoylated, we examined the importance of interacting with the G protein betagamma dimer for their proper plasma membrane localization and palmitoylation. Conserved alpha subunit N-terminal amino acids predicted to mediate binding to betagamma were mutated to create a series of betagamma binding region mutants expressed in HEK293 cells. These alpha(s) and alpha(q) mutants were found in soluble rather than particulate fractions, and they no longer localized to plasma membranes as demonstrated by immunofluorescence microscopy. The mutations also inhibited incorporation of radiolabeled palmitate into the proteins and abrogated their signaling ability. Additional alpha(q) mutants, which contain these mutations but are modified by both myristate and palmitate, retained their localization to plasma membranes and ability to undergo palmitoylation. These findings identify binding to betagamma as a critical membrane attachment signal for alpha(s) and alpha(q) and as a prerequisite for their palmitoylation, while myristoylation can restore membrane localization and palmitoylation of betagamma binding-deficient alpha(q) subunits.
Signal transduction via G-protein-coupled receptors (GPCRs) is a fundamental pathway through which the functions of an individual cell can be integrated within the demands of a multicellular organism. Since this family of receptors first discovered, the proteins that constitute this signaling cascade and their interactions with one another have been studied intensely. In parallel, the pivotal role of lipids in the correct and efficient propagation of extracellular signals has attracted ever increasing attention. This is not surprising given that most of the signal transduction machinery is membrane-associated and therefore lipid-related. Hence, lipid-protein interactions exert a considerable influence on the activity of these proteins. This review focuses on the post-translational lipid modifications of GPCRs and G proteins (palmitoylation, myristoylation, and isoprenylation) and their significance for membrane binding, trafficking and signaling. Moreover, we address how the particular biophysical properties of different membrane structures may regulate the localization of these proteins and the potential functional consequences of this phenomenon in signal transduction. Finally, the interactions that occur between membrane lipids and GPCR effector enzymes such as PLC and PKC are also considered.
We have examined the subcellular distribution of as/ the a subunit of the heterotrimeric G protein Gs, by using INTRODUCTIONRegulated and reversible translocation within the cell is a common paradigm for proteins involved in signal transduction pathways. Upon extracellular stimulation, many such proteins reversibly translocate from the plasma membrane to cytoplasm, from cytoplasm to the plasma membrane or nucleus, or between different membrane locations.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.