An approach to the study of G-protein-coupled receptor kinases: an in vitro-purified membrane assay reveals differential receptor specificity and regulation by G beta gamma subunits.

TNFα (tumour necrosis factor α) is a multifunctional cytokine involved in the pathophysiology of many chronic inflammatory diseases. TNFα activation of the NF-κB (nuclear factor κB) signalling pathway particularly in macrophages has been implicated in many diseases. We demonstrate in the present study that GRK2 and GRK5 (G-protein-coupled-receptor kinases 2 and 5) regulate TNFα-induced NF-κB signalling in Raw 264.7 macrophages. RNAi (RNA interference) knockdown of GRK2 or GRK5 in macrophages significantly inhibited TNFα-induced IκBα (inhibitory κBα) phosphorylation and degradation, NF-κB activation and expression of the NF-κB-regulated gene MIP1β (macrophage inflammatory protein 1β). Consistent with these results, overexpression of GRK2 or GRK5 enhanced TNFα-induced NF-κB activity. In addition, we show that GRK2 and GRK5 interacted with IκBα via the N-terminal domain of IκBα and that IκBα is a substrate for GRK2 and GRK5 in vitro. Furthermore, we also found that GRK5, but not GRK2, phosphorylated IκBα at the same amino acid residues (Ser32/Ser36) as that of IKKβ (IκB kinase β). Interestingly, associated with these results, knockdown of IKKβ in Raw 264.7 macrophages did not affect TNFα-induced IκBα phosphorylation. Taken together, these results demonstrate that both GRK2 and GRK5 are important and novel mediators of a non-traditional IκBα/NF-κB signalling pathway.

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Receptor specificity of G-protein-coupled receptor kinases in target cells

Trends in Pharmacological Sciences ◽

10.1016/s0165-6147(00)01641-2 ◽

2001 ◽

Vol 22 (4) ◽

pp. 168-169 ◽

Cited By ~ 1

Author(s):

Michele Sallese ◽

Luisa Iacovelli ◽

Marianna Storto ◽

Antonio De Blasi

Keyword(s):

G Protein ◽

Target Cells ◽

G Protein Coupled Receptor ◽

Receptor Specificity ◽

Receptor Kinases ◽

G Protein Coupled

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GRK Inhibition Potentiates Glucagon-Like Peptide-1 Action

Frontiers in Endocrinology ◽

10.3389/fendo.2021.652628 ◽

2021 ◽

Vol 12 ◽

Author(s):

Seunghun P. Lee ◽

Jenson Qi ◽

Guozhang Xu ◽

Matthew M. Rankin ◽

James Littrell ◽

...

Keyword(s):

G Protein ◽

G Protein Coupled Receptor ◽

Dipeptidyl Peptidase 4 ◽

Receptor Kinases ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1 ◽

Receptor Interacting Proteins ◽

G Protein Coupled

The glucagon-like peptide-1 receptor (GLP-1R) is a G-protein-coupled receptor (GPCR) whose activation results in suppression of food intake and improvement of glucose metabolism. Several receptor interacting proteins regulate the signaling of GLP-1R such as G protein-coupled receptor kinases (GRK) and β-arrestins. Here we evaluated the physiological and pharmacological impact of GRK inhibition on GLP-1R activity leveraging small molecule inhibitors of GRK2 and GRK3. We demonstrated that inhibition of GRK: i) inhibited GLP-1-mediated β-arrestin recruitment, ii) enhanced GLP-1-induced insulin secretion in isolated islets and iii) has additive effect with dipeptidyl peptidase 4 in mediating suppression of glucose excursion in mice. These findings highlight the importance of GRK to modulate GLP-1R function in vitro and in vivo. GRK inhibition is a potential therapeutic approach to enhance endogenous and pharmacologically stimulated GLP-1R signaling.

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Increased expression of G-protein-coupled receptor kinases 3 and 4 in hyperfunctioning thyroid nodules

Journal of Endocrinology ◽

10.1677/joe.0.1820173 ◽

2004 ◽

Vol 182 (1) ◽

pp. 173-182 ◽

Cited By ~ 16

Author(s):

C Voigt ◽

HP Holzapfel ◽

S Meyer ◽

R Paschke

Keyword(s):

G Protein ◽

Thyroid Nodules ◽

Thyroid Tissue ◽

Human Thyroid ◽

G Protein Coupled Receptor ◽

Hek 293 ◽

293 Cells ◽

Receptor Kinases ◽

G Protein Coupled

G-protein-coupled receptor kinases (GRKs) are implicated in the pathophysiology of human diseases such as arterial hypertension, heart failure and rheumatoid arthritis. While G-protein-coupled receptor kinases 2 and 5 have been shown to be involved in the desensitization of the rat thyrotropin receptor (TSHR), their role in the pathophysiology of hyperfunctioning thyroid nodules (HTNs) is unknown. Therefore, we analyzed the expression pattern of the known GRKs in human thyroid tissue and investigated their function in the pathology of HTNs. The expression of different GRKs in human thyroid and HTNs was measured by Western blotting. The influence of GRK expression on TSHR function was analyzed by coexpression experiments in HEK 293 cells. We demonstrate that in addition to GRKs 2, 5 and 6, GRKs 3 and 4 are also expressed in the human thyroid. GRKs 2, 3, 5 and 6 are able to desensitize the TSHR in vitro. This GRK-induced desensitization is amplified by the additional over-expression of beta-arrestin 1 or 2. We did not find any mutations in the GRKs 2, 3 and 5 from 14 HTNs without TSHR mutations and Gsalpha mutations. The expression of GRKs 3 and 4 was increased in HTNs independently from the existence of TSHR mutations or Gsalpha mutations. In conclusion, the increased expression of GRK 3 in HTNs and the ability of GRK 3 to desensitize the TSHR in vitro, suggest a potential role for GRK 3 as a negative feedback regulator for the constitutively activated cAMP pathway in HTNs.

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Reply: receptor specificity of G-protein-coupled receptor kinases

Trends in Pharmacological Sciences ◽

10.1016/s0165-6147(00)01538-8 ◽

2000 ◽

Vol 21 (10) ◽

pp. 366-367 ◽

Cited By ~ 22

Author(s):

Raul R Gainetdinov ◽

Richard T Premont ◽

Marc G Caron ◽

Robert J Lefkowitz

Keyword(s):

G Protein ◽

G Protein Coupled Receptor ◽

Receptor Specificity ◽

Receptor Kinases ◽

G Protein Coupled

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The G Protein-Coupled Receptor Kinases (GRKs) in Chemokine Receptor-Mediated Immune Cell Migration: From Molecular Cues to Physiopathology

Cells ◽

10.3390/cells10010075 ◽

2021 ◽

Vol 10 (1) ◽

pp. 75

Author(s):

Marta Laganà ◽

Géraldine Schlecht-Louf ◽

Françoise Bachelerie

Keyword(s):

G Protein ◽

Chemokine Receptor ◽

Immune Cell ◽

Neutrophil Migration ◽

G Protein Coupled Receptor ◽

Receptor Kinases ◽

Immune Cell Migration ◽

And Migration ◽

G Protein Coupled ◽

Gpcr Desensitization

Although G protein-coupled receptor kinases (GRKs) have long been known to regulate G protein-coupled receptor (GPCR) desensitization, their more recently characterized functions as scaffolds and signalling adapters underscore that this small family of proteins governs a larger array of physiological functions than originally suspected. This review explores how GRKs contribute to the complex signalling networks involved in the migration of immune cells along chemokine gradients sensed by cell surface GPCRs. We outline emerging evidence indicating that the coordinated docking of several GRKs on an active chemokine receptor determines a specific receptor phosphorylation barcode that will translate into distinct signalling and migration outcomes. The guidance cues for neutrophil migration are emphasized based on several alterations affecting GRKs or GPCRs reported to be involved in pathological conditions.

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