Spatial regulation of GPR64/ADGRG2 signaling by β‐arrestins and GPCR kinases

Azimzadeh, Pedram; Talamantez-Lyburn, Sarah C.; Chang, Katarina T.; Inoue, Asuka; Balenga, Nariman

doi:10.1111/nyas.14227

Cited by 19 publications

(36 citation statements)

References 48 publications

(73 reference statements)

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“…By applying our newly developed agonist VPM-p15, we further demonstrated that the arrestins could be recruited to ADGRG2 in an agonist-dependent manner. The small differences between previous results and ours regarding the p15-induced arrestin recruitment to wild-type ADGRG2 may be due to altered experimental methods with different sensitivities (24).…”

Section: Discussioncontrasting

confidence: 85%

“…In this study, we found that the ADGRG2-β (CTF fragment) showed higher constitutive activity for arrestin recruitment compared to that of the full-length ADGRG2 (Supplemental Figure 1E-F). Consistently, a recent study reported that △NTF and P622 mutants of ADGRG2 interacted with β-arrestin1 and β-arrestin2, which induced constitutive internalization of ADGRG2 in steady states (24). Therefore, the arrestin recruitment to ADGRG2 might be strengthened by activation of the ADGRG2 via Stachel sequence interaction.…”

Section: Discussionsupporting

confidence: 67%

“…In our recent studies, we have found that ADGRG2 is specifically coupled to the cystic fibrosis transmembrane conductance regulator (CFTR) through a Gq-and β-arrestin1-mediated mechanism, regulating the ion/water homeostasis in efferent ductules which is essential for male fertility. It has been shown that ADGRG2 could be activated by the Stachel peptide p15(sequence: TSFGILLDLSRTSLP) derived from the Stachel sequence(23) ， to elicit downstream Gs, Gq, G12/13 signaling ( Figure 1A) (23)(24)(25). p15 does not activate or control other aGPCRs (GPR110, GPR133), thereby displaying apparent specificities for the receptor it originated from (23).…”

Section: Introductionmentioning

confidence: 99%

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Optimization of a peptide ligand for the adhesion GPCR ADGRG2 provides a potent tool to explore receptor biology

Sun

Zhang

et al. 2021

Journal of Biological Chemistry

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The adhesion GPCR ADGRG2, also known as GPR64, is a critical regulator of male fertility that maintains ion/pH homeostasis and CFTR coupling. The molecular basis of ADGRG2 function is poorly understood, in part because no endogenous ligands for ADGRG2 have been reported, thus limiting the tools available to interrogate ADGRG2 activity. It has been shown that ADGRG2 can be activated by a peptide, termed p15, derived from its own N-terminal region known as the Stachel sequence. However, the low affinity of p15 limits its utility for ADGRG2 characterization. In the current study, we used alanine scanning mutagenesis to examine the critical residues responsible for p15-induced ADGRG2 activity. We next designed systematic strategies to optimize the peptide agonist of ADGRG2, using natural and unnatural amino acid substitutions. We obtained an optimized ADGRG2 Stachel peptide T1V/F3Phe(4-Me) (VPM-p15) that activated ADGRG2 with significantly improved (>2 orders of magnitude) affinity. We then characterized the residues in ADGRG2 that were important for ADGRG2 activation in response to VPM-p15 engagement, finding that the toggle switch W 6.53 and residues of the ECL2 region of ADGRG2 are key determinants for VPM-p15 interactions and VPM-p15-induced Gs or arrestin signaling. Our study not only provides a useful tool to investigate the function of ADGRG2 but also offers new insights to guide further optimization of Stachel peptides to activate adhesion GPCR members.

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Section: Discussioncontrasting

confidence: 85%

Section: Discussionsupporting

confidence: 67%

Section: Introductionmentioning

confidence: 99%

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Optimization of a peptide ligand for the adhesion GPCR ADGRG2 provides a potent tool to explore receptor biology

Sun

Zhang

et al. 2021

Journal of Biological Chemistry

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“…It was found that the TM1 N-terminal stalk sequence consisting of ~20 residues behaved as a tethered-peptide-agonist and dramatically activated AGPCRs. Deletion of tethered agonist residues dramatically lowered receptor activities in vitro, as shown for ADGRG1, ADGRG2 (GPR64), ADGRG6, and ADGRD1 (GPR133), and ADGRF1 (21,22,40,41). Partial deletion of the zebrafish ADGRG6 tethered agonist resulted in a puffy ear phenotype and impairment of nerve cell myelination, essentially phenocopying the fish model deletion of the ADGRG6 CTF (22,42).…”

Section: Agonism (Tethered-peptide Agonism)mentioning

confidence: 73%

Mechanisms of adhesion G protein–coupled receptor activation

Vizurraga

Adhikari

Yeung

et al. 2020

Journal of Biological Chemistry

115

141

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Adhesion G protein coupled receptors (AGPCRs) are a thirty-three-member subfamily of Class B GPCRs that control a wide array of physiological processes and are implicated in disease. AGPCRs uniquely contain large, self-proteolyzing extracellular regions that range from hundreds to thousands of residues in length. AGPCR autoproteolysis occurs within the extracellular GPCR Autoproteolysis-Inducing (GAIN) domain that is proximal to the N-terminus of the G protein-coupling seven transmembrane spanning (7TM) bundle. GAIN domain-mediated self-cleavage is constitutive and produces two-fragment holoreceptors that remain bound at the cell surface. It has been of recent interest to understand how AGPCRs are activated in relation to their two-fragment topologies. Dissociation of the AGPCR fragments stimulates G protein signaling through the action of the tethered-peptide-agonist stalk that is occluded within the GAIN domain in the holoreceptor form. AGPCRs can also signal independently of fragment dissociation, and a few receptors possess GAIN domains incapable of self-proteolysis. This has resulted in complex theories as to how these receptors are activated in vivo, complicating pharmacological advances. Currently there is no existing structure of an activated AGPCR to support any of the theories. Further confounding AGPCR research is that many of the receptors remain orphans and lack identified activating ligands. In this review, we provide a detailed layout of the current theorized modes of AGPCR activation with discussion of potential parallels to mechanisms used by other GPCR classes. We provide a classification means for the ligands that have been identified and discuss how these ligands may activate AGPCRs in physiological-contexts.

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“…These studies established ADGRG2 as a novel gene responsible for a specific subtype of obstructive azoospermia. The molecular mechanism of X-linked CBAVD remains uncertain, although a regulation of fluid reabsorption by G-protein-dependent CFTR coupling and β-arrestin-dependent receptor internalization has been postulated (Zhang et al 2018 ; Azimzadeh et al 2019 ).…”

Section: Obstructive Azoospermiamentioning

confidence: 99%

The X chromosome and male infertility

et al. 2019

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The X chromosome is a key player in germ cell development, as has been highlighted for males in previous studies revealing that the mammalian X chromosome is enriched in genes expressed in early spermatogenesis. In this review, we focus on the X chromosome's unique biology as associated with human male infertility. Male infertility is most commonly caused by spermatogenic defects to which X chromosome dosage is closely linked; for example, any supernumerary X chromosome as in Klinefelter syndrome will lead to male infertility. Furthermore, because males normally only have a single X chromosome and because X-linked genetic anomalies are generally only present in a single copy in males, any loss-of-function mutations in single-copy X-chromosomal genes cannot be compensated by a normal allele. These features make X-linked genes particularly attractive for studying male spermatogenic failure. However, to date, only very few genetic causes have been identified as being definitively responsible for male infertility in humans. Although genetic studies of germ cell-enriched X-chromosomal genes in mice suggest a role of certain human orthologs in infertile men, these genes in mice and humans have striking evolutionary differences. Furthermore, the complexity and highly repetitive structure of the X chromosome hinder the mutational analysis of X-linked genes in humans. Therefore, we conclude that additional methodological approaches are urgently warranted to advance our understanding of the genetics of X-linked male infertility.Matthias Vockel and Antoni Riera-Escamilla are co-first authors.Frank Tüttelmann and Csilla Krausz are co-last authors.

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Spatial regulation of GPR64/ADGRG2 signaling by β‐arrestins and GPCR kinases

Cited by 19 publications

References 48 publications

Optimization of a peptide ligand for the adhesion GPCR ADGRG2 provides a potent tool to explore receptor biology

Optimization of a peptide ligand for the adhesion GPCR ADGRG2 provides a potent tool to explore receptor biology

Mechanisms of adhesion G protein–coupled receptor activation

The X chromosome and male infertility

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