scholarly journals A short C-terminal peptide in Gγ regulates Gβγ signaling efficacy

2021 ◽  
pp. mbc.E20-11-0750
Author(s):  
Mithila Tennakoon ◽  
Kanishka Senarath ◽  
Dinesh Kankanamge ◽  
Deborah N. Chadee ◽  
Ajith Karunarathne
Keyword(s):  
Amino Acids ◽  
Plasma Membrane ◽  
G Protein ◽  
G Protein Signaling ◽  
Protein Signaling ◽  
Diverse Range ◽  
Cell Behaviors ◽  
Molecular Process ◽  
Carboxy Terminal ◽  
Molecular Picture

G protein beta-gamma (Gβγ) subunits anchor to the plasma membrane (PM) through the carboxy-terminal (CT) prenyl group in Gγ. This interaction is crucial for the PM localization and functioning of Gβγ, allowing GPCR-G protein signaling to proceed. The diverse Gγ family has twelve members, and we have recently shown that the signaling efficacies of major Gβγ effectors are Gγ-type dependent. This dependency is due to the distinct series of membrane-interacting abilities of Gγ. However, the molecular process allowing for Gβγ subunits to exhibit a discrete and diverse range of Gγ-type dependent membrane affinities is unclear and cannot be explained only using the type of prenylation. The present work explores the unique designs of membrane-interacting CT-residues in Gγ as a major source for this Gγ-type dependent Gβγ signaling. Despite the type of prenylation, results show signaling efficacy at the PM, and associated cell behaviors of Gβγ are governed by crucially located specific amino acids in the 5–6 residue pre-prenylation region of Gγ. The provided molecular picture of Gγ–membrane interactions may explain how cells gain Gγ-type dependent G protein-GPCR signaling as well as how Gβγ elicits selective signaling at various subcellular compartments.

scholarly journals Subcellular optogenetic inhibition of G proteins generates signaling gradients and cell migration

2014 ◽  
Vol 25 (15) ◽  
pp. 2305-2314 ◽  
Author(s):  
Patrick R. O'Neill ◽  
N. Gautam
Keyword(s):  
Cell Migration ◽  
G Protein ◽  
Intracellular Signaling ◽  
Signaling Molecules ◽  
G Protein Signaling ◽  
Protein Signaling ◽  
Cell Behaviors ◽  
Directed Cell Migration ◽  
Neurite Initiation ◽  
Mechanistic Basis

Cells sense gradients of extracellular cues and generate polarized responses such as cell migration and neurite initiation. There is static information on the intracellular signaling molecules involved in these responses, but how they dynamically orchestrate polarized cell behaviors is not well understood. A limitation has been the lack of methods to exert spatial and temporal control over specific signaling molecules inside a living cell. Here we introduce optogenetic tools that act downstream of native G protein–coupled receptor (GPCRs) and provide direct control over the activity of endogenous heterotrimeric G protein subunits. Light-triggered recruitment of a truncated regulator of G protein signaling (RGS) protein or a Gβγ-sequestering domain to a selected region on the plasma membrane results in localized inhibition of G protein signaling. In immune cells exposed to spatially uniform chemoattractants, these optogenetic tools allow us to create reversible gradients of signaling activity. Migratory responses generated by this approach show that a gradient of active G protein αi and βγ subunits is sufficient to generate directed cell migration. They also provide the most direct evidence so for a global inhibition pathway triggered by Gi signaling in directional sensing and adaptation. These optogenetic tools can be applied to interrogate the mechanistic basis of other GPCR-modulated cellular functions.

scholarly journals The regulator of G protein signaling (RGS) domain of G protein–coupled receptor kinase 5 (GRK5) regulates plasma membrane localization and function

2014 ◽  
Vol 25 (13) ◽  
pp. 2105-2115 ◽  
Author(s):  
Hua Xu ◽  
Xiaoshan Jiang ◽  
Ke Shen ◽  
Christopher C. Fischer ◽  
Philip B. Wedegaertner
Keyword(s):  
Plasma Membrane ◽  
G Protein ◽  
Calcium Release ◽  
Membrane Localization ◽  
G Protein Signaling ◽  
Protein Signaling ◽  
G Protein Coupled Receptor ◽  
Rgs Domain ◽  
And Function ◽  
G Protein Coupled

The G protein–coupled receptor (GPCR) kinases (GRKs) phosphorylate activated GPCRs at the plasma membrane (PM). Here GRK5/GRK4 chimeras and point mutations in GRK5 identify a short sequence within the regulator of G protein signaling (RGS) domain in GRK5 that is critical for GRK5 PM localization. This region of the RGS domain of GRK5 coincides with a region of GRK6 and GRK1 shown to form a hydrophobic dimeric interface (HDI) in crystal structures. Coimmunoprecipitation (coIP) and acceptor photobleaching fluorescence resonance energy transfer assays show that expressed GRK5 self-associates in cells, whereas GRK5-M165E/F166E (GRK5-EE), containing hydrophilic mutations in the HDI region of the RGS domain, displays greatly decreased coIP interactions. Both forcing dimerization of GRK5-EE, via fusion to leucine zipper motifs, and appending an extra C-terminal membrane-binding region to GRK5-EE (GRK5-EE-CT) recover PM localization. In addition, GRK5-EE displays a decreased ability to inhibit PAR1-induced calcium release compared with GRK5 wild type (wt). In contrast, PM-localized GRK5-EE-CaaX (appending a C-terminal prenylation and polybasic motif from K-ras) or GRK5-EE-CT shows comparable ability to GRK5 wt to inhibit PAR1-induced calcium release. The results suggest a novel model in which GRK5 dimerization is important for its plasma membrane localization and function.

G protein Signaling, Journeys Beyond the Plasma Membrane

2017 ◽  
Vol 97 (1) ◽  
pp. 95-108 ◽  
Author(s):  
Raji R. Nair ◽  
Amritanjali Kiran ◽  
Deepak Kumar Saini
Keyword(s):  
Plasma Membrane ◽  
G Protein ◽  
G Protein Signaling ◽  
Protein Signaling

scholarly journals RSBP-1 Is a Membrane-targeting Subunit Required by the Gαq-specific But Not the Gαo-specific R7 Regulator of G protein Signaling in Caenorhabditis elegans

2010 ◽  
Vol 21 (2) ◽  
pp. 232-243 ◽  
Author(s):  
Morwenna Y. Porter ◽  
Michael R. Koelle
Keyword(s):  
Plasma Membrane ◽  
Caenorhabditis Elegans ◽  
G Protein ◽  
Cultured Cells ◽  
Membrane Targeting ◽  
G Protein Signaling ◽  
Rgs Proteins ◽  
Protein Signaling ◽  
Functional Requirements ◽  
Rgs Protein

Regulator of G protein signaling (RGS) proteins inhibit G protein signaling by activating Gα GTPase activity, but the mechanisms that regulate RGS activity are not well understood. The mammalian R7 binding protein (R7BP) can interact with all members of the R7 family of RGS proteins, and palmitoylation of R7BP can target R7 RGS proteins to the plasma membrane in cultured cells. However, whether endogenous R7 RGS proteins in neurons require R7BP or membrane localization for function remains unclear. We have identified and knocked out the only apparent R7BP homolog in Caenorhabditis elegans, RSBP-1. Genetic studies show that loss of RSBP-1 phenocopies loss of the R7 RGS protein EAT-16, but does not disrupt function of the related R7 RGS protein EGL-10. Biochemical analyses find that EAT-16 coimmunoprecipitates with RSBP-1 and is predominantly plasma membrane-associated, whereas EGL-10 does not coimmunoprecipitate with RSBP-1 and is not predominantly membrane-associated. Mutating the conserved membrane-targeting sequence in RSBP-1 disrupts both the membrane association and function of EAT-16, demonstrating that membrane targeting by RSBP-1 is essential for EAT-16 activity. Our analysis of endogenous R7 RGS proteins in C. elegans neurons reveals key differences in the functional requirements for membrane targeting between members of this protein family.

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