CaaX-motif-adjacent residues influence G protein gamma (Gγ) prenylation under suboptimal conditions

Tennakoon, Mithila; Thotamune, Waruna; Payton, John L.; Karunarathne, Ajith

doi:10.1016/j.jbc.2023.105269

Cited by 3 publications

(2 citation statements)

References 87 publications

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“…Building on our previous work highlighting the substantial involvement of free Gβγ in PIP2 hydrolysis and its attenuation ( 10 ), we here demonstrated a negative impact on the canonical PIP2 hydrolysis attenuation observed upon Gq-GPCR activation through the OptoGDI-induced optogenetic release of free Gβγ. Using a Gγ subunit that provides the Gβγ complex the lowest observed membrane affinity, Gγ9 ( 53 , 54 ), we show the reversible Gβγ release in single cells and from subcellular regions. To our understanding, this is the first direct demonstration of real-time GDI signaling in living cells.…”

Section: Discussionmentioning

confidence: 96%

AGS3-based optogenetic GDI induces GPCR-independent Gβγ signaling and macrophage migration

Thotamune,

Ubeysinghe,

Rajarathna

et al. 2024

Preprint

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G protein-coupled receptors (GPCRs) are efficient Guanine nucleotide exchange factors (GEFs) and exchange GDP to GTP on the Gα subunit of G protein heterotrimers in response to various extracellular stimuli, including neurotransmitters and light. GPCRs primarily broadcast signals through activated G proteins, GαGTP, and free Gβγ and are major disease drivers. Evidence shows that the ambient low threshold signaling required for cells is likely supplemented by signaling regulators such as non-GPCR GEFs and Guanine nucleotide Dissociation Inhibitors (GDIs). Activators of G protein Signaling 3 (AGS3) are recognized as a GDI involved in multiple health and disease-related processes. Nevertheless, understanding of AGS3 is limited, and no significant information is available on its structure-function relationship or signaling regulation in living cells. Here, we employedin silicostructure-guided engineering of a novel optogenetic GDI, based on the AGS3’s G protein regulatory (GPR) motif, to understand its GDI activity and induce standalone Gβγ signaling in living cells on optical command. Our results demonstrate that plasma membrane recruitment of OptoGDI efficiently releases Gβγ, and its subcellular targeting generated localized PIP3 and triggered macrophage migration. Therefore, we propose OptoGDI as a powerful tool for optically dissecting GDI-mediated signaling pathways and triggering GPCR-independent Gβγ signaling in cells andin vivo.

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Section: Discussionmentioning

confidence: 96%

AGS3-based optogenetic GDI induces GPCR-independent Gβγ signaling and macrophage migration

Thotamune,

Ubeysinghe,

Rajarathna

et al. 2024

Preprint

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“…G proteins heterotrimers contain 2-3 lipid modifications: myristoyl or palmitoyl, or both on the N-terminus of G and prenyl modification on the C-terminus of G. 62 Therefore, mobility of heterotrimers through the cytosol is not favored, and the lateral diffusion through the membranes is a relatively slow process. On the contrary, free G shuttles efficiently through the cytosol in a G subtype dependent manner, in which G9 provided the G the fastest shuttling rate.…”

Section: Optoiso Induced Endomembrane Exclusive 1ar Signalingmentioning

confidence: 99%

Optical Control of Cell-Surface and Endomembrane-Exclusive β-Adrenergic Receptor Signaling

Thotamune,

Ubeysinghe,

Shrestha

et al. 2024

Preprint

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Beta-adrenergic receptors (βARs) are G protein-coupled receptors (GPCRs) that mediate catecholamine-induced stress responses, such as heart rate increase and bronchodilation. In addition to signals from the cell surface, βARs also broadcast non-canonical signaling activities from the cell interior membranes (endomembranes). Dysregulation of these receptor pathways underlies severe pathological conditions. Excessive βAR stimulation is linked to cardiac hypertrophy, leading to heart failure, while impaired stimulation causes compromised fight or flight stress responses and homeostasis. In addition to plasma membrane βAR, emerging evidence indicates potential pathological implications of deeper endomembrane βARs, such as inducing cardiomyocyte hypertrophy and apoptosis, underlying heart failure. However, the lack of approaches to control their signaling in subcellular compartments exclusively has impeded linking endomembrane βAR signaling with pathology. Informed by the β1AR-catecholamine interactions, we engineered an efficiently photo-labile, protected hydroxy β1AR pro-ligand (OptoIso) to trigger βAR signaling at the cell surface, as well as exclusive endomembrane regions upon blue light stimulation. Not only does OptoIso undergo blue light deprotection in seconds, but it also efficiently enters cells and allows examination of G protein heterotrimer activation exclusively at endomembranes. In addition to its application in the optical interrogation of βARs in unmodified cells, given its ability to control deep organelle βAR signaling, OptoIso will be a valuable experimental tool.

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Optical control of cell-surface and endomembrane-exclusive β-adrenergic receptor signaling

Thotamune,

Ubeysinghe,

Shrestha

et al. 2024

Journal of Biological Chemistry

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CaaX-motif-adjacent residues influence G protein gamma (Gγ) prenylation under suboptimal conditions

Cited by 3 publications

References 87 publications

AGS3-based optogenetic GDI induces GPCR-independent Gβγ signaling and macrophage migration

AGS3-based optogenetic GDI induces GPCR-independent Gβγ signaling and macrophage migration

Optical Control of Cell-Surface and Endomembrane-Exclusive β-Adrenergic Receptor Signaling

Optical control of cell-surface and endomembrane-exclusive β-adrenergic receptor signaling

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