scholarly journals Discovery of a dual Ras and ARF6 inhibitor from a GPCR endocytosis screen

2020 ◽  
Author(s):  
Jenna Giubilaro ◽  
Doris Schuetz ◽  
Yoon Namkung ◽  
Etienne Khoury ◽  
Monica Marquez ◽  
...  
Keyword(s):  
G Protein ◽  
Tyrosine Kinases ◽  
Hormone Receptors ◽  
Receptor Trafficking ◽  
Cancer Cell Proliferation ◽  
New Class ◽  
In Silico Modeling ◽  
G Protein Coupled

Abstract Internalization and intracellular trafficking of hormone receptors, like receptor tyrosine kinases (RTKs) and G protein-coupled receptors (GPCRs), play pivotal roles in cell responsiveness homeostasis. Dysregulation in receptor trafficking can lead to aberrant signaling and cell behavior, prevalent in cancer. Here, using an endosomal BRET-based assay in a high-throughput screen with the prototypical GPCR angiotensin II type 1 receptor (AT1R), we sought to identify inhibitors of receptor trafficking from a library of ~115,000 small molecules. We identified a novel dual Ras and ARF6 inhibitor that blocks agonist-mediated internalization of AT1R and other GPCRs, which we named Rasarfin. Rasarfin also potently inhibited agonist-induced ERK1/2 signaling by GPCRs, and MAPK and Akt signaling by EGFR, as well as prevented cancer cell proliferation. In silico modeling and in vitro studies revealed a unique binding modality of Rasarfin within the SOS-binding domain of Ras. Our findings unveil a new class of dual small G protein inhibitors for receptor trafficking and signaling, useful for the inhibition of oncogenic cellular responses.

scholarly journals Discovery of a dual Ras and ARF6 inhibitor from a GPCR endocytosis screen

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jenna Giubilaro ◽  
Doris A. Schuetz ◽  
Tomasz M. Stepniewski ◽  
Yoon Namkung ◽  
Etienne Khoury ◽  
...  
Keyword(s):  
G Protein ◽  
Cellular Responses ◽  
Receptor Trafficking ◽  
G Protein Coupled Receptors ◽  
Cancer Cell Proliferation ◽  
Small G Protein ◽  
In Silico Modeling ◽  
G Protein Coupled

AbstractInternalization and intracellular trafficking of G protein-coupled receptors (GPCRs) play pivotal roles in cell responsiveness. Dysregulation in receptor trafficking can lead to aberrant signaling and cell behavior. Here, using an endosomal BRET-based assay in a high-throughput screen with the prototypical GPCR angiotensin II type 1 receptor (AT1R), we sought to identify receptor trafficking inhibitors from a library of ~115,000 small molecules. We identified a novel dual Ras and ARF6 inhibitor, which we named Rasarfin, that blocks agonist-mediated internalization of AT1R and other GPCRs. Rasarfin also potently inhibits agonist-induced ERK1/2 signaling by GPCRs, and MAPK and Akt signaling by EGFR, as well as prevents cancer cell proliferation. In silico modeling and in vitro studies reveal a unique binding modality of Rasarfin within the SOS-binding domain of Ras. Our findings unveil a class of dual small G protein inhibitors for receptor trafficking and signaling, useful for the inhibition of oncogenic cellular responses.

scholarly journals G Protein-Coupled Estrogen Receptor 1 (GPER) as a Novel Target for Schizophrenia Drug Treatment

2020 ◽  
Vol 1 (1) ◽  
Author(s):  
Danielle S Macêdo ◽  
Lia Lira Olivier Sanders ◽  
Raimunda das Candeias ◽  
Cyntia de Freitas Montenegro ◽  
David Freitas de Lucena ◽  
...  
Keyword(s):  
Estrogen Receptor ◽  
Estrogen Receptors ◽  
G Protein ◽  
Brain Inflammation ◽  
Antipsychotic Treatment ◽  
New Class ◽  
Estrogen Receptor Modulators ◽  
Novel Target ◽  
G Protein Coupled ◽  
Estrogen Receptor 1

Abstract The observation that a person’s sex influences the onset age of schizophrenia, the course of the disease, and antipsychotic treatment response suggests a possible role for estrogen receptors in the pathophysiology of schizophrenia. Indeed, treatment with adjunctive estrogen or selective estrogen receptor modulators (SERMs) are known to reduce schizophrenia symptoms. While estrogen receptors (ER)α and ERβ have been studied, a third and more recently discovered estrogen receptor, the G protein-coupled estrogen receptor 1 (GPER), has been largely neglected. GPER is a membrane receptor that regulates non-genomic estrogen functions, such as the modulation of emotion and inflammatory response. This review discusses the possible role of GPER in brain impairments seen in schizophrenia and in its potential as a therapeutic target. We conducted a comprehensive literature search in the PubMed/MEDLINE database, using the following search terms: “Schizophrenia,” “Psychosis,” “GPER1 protein,” “Estrogen receptors,” “SERMS,” “GPER1 agonism, “Behavioral symptoms,” “Brain Inflammation.” Studies involving GPER in schizophrenia, whether preclinical or human studies, have been scarce, but the results are encouraging. Agonism of the GPER receptor could prove to be an essential mechanism of action for a new class of “anti-schizophrenia” drugs.

scholarly journals Adhesion G Protein–Coupled Receptors: From In Vitro Pharmacology to In Vivo Mechanisms

2015 ◽  
Vol 88 (3) ◽  
pp. 617-623 ◽  
Author(s):  
Kelly R. Monk ◽  
Jörg Hamann ◽  
Tobias Langenhan ◽  
Saskia Nijmeijer ◽  
Torsten Schöneberg ◽  
...  
Keyword(s):  
G Protein ◽  
G Protein Coupled Receptors ◽  
In Vitro Pharmacology ◽  
G Protein Coupled

scholarly journals The treatment effects of flaxseed-derived secoisolariciresinol diglycoside and its metabolite enterolactone on benign prostatic hyperplasia involve the G protein-coupled estrogen receptor 1

2016 ◽  
Vol 41 (12) ◽  
pp. 1303-1310 ◽  
Author(s):  
Guan-Yu Ren ◽  
Chun-Yang Chen ◽  
Wei-Guo Chen ◽  
Ya Huang ◽  
Li-Qiang Qin ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Estrogen Receptor ◽  
Benign Prostatic Hyperplasia ◽  
G Protein ◽  
Therapeutic Potential ◽  
Prostatic Hyperplasia ◽  
Docking Simulations ◽  
G Protein Coupled

Secoisolariciresinol diglucoside (SDG), a lignan extracted from flaxseed, has been shown to suppress benign prostatic hyperplasia (BPH). However, little is known about the mechanistic basis for its anti-BPH activity. The present study showed that enterolactone (ENL), the mammalian metabolite of SDG, shared the similar binding site of G1 on a new type of membranous estrogen receptor, G-protein-coupled estrogen eceptor 1 (GPER), by docking simulations method. ENL and G1 (the specific agonist of GPER) inhibited the proliferation of human prostate stromal cell line WPMY-1 as shown by MTT assay and arrested cell cycle at the G0/G1 phase, which was displayed by propidium iodide staining following flow cytometer examination. Silencing GPER by short interfering RNA attenuated the inhibitory effect of ENL on WPMY-1 cells. The therapeutic potential of SDG in the treatment of BPH was confirmed in a testosterone propionate-induced BPH rat model. SDG significantly reduced the enlargement of the rat prostate and the number of papillary projections of prostatic alveolus and thickness of the pseudostratified epithelial and stromal cells when comparing with the model group. Mechanistic studies showed that SDG and ENL increased the expression of GPER both in vitro and in vivo. Furthermore, ENL-induced cell cycle arrest may be mediated by the activation of GPER/ERK pathway and subsequent upregulation of p53 and p21 and downregulation of cyclin D1. This work, in tandem with previous studies, will enhance our knowledge regarding the mechanism(s) of dietary phytochemicals on BPH prevention and ultimately expand the scope of adopting alternative approaches in BPH treatment.

scholarly journals Characterization of Two Fly LGR (Leucine-Rich Repeat-Containing, G Protein-Coupled Receptor) Proteins Homologous to Vertebrate Glycoprotein Hormone Receptors: Constitutive Activation of Wild-Type Fly LGR1 But Not LGR2 in Transfected Mammalian Cells**This study was supported by NIH Grant HD-23273. The GenBank submission number for fly LGR2 is AF274591.

Endocrinology ◽  
2000 ◽  
Vol 141 (11) ◽  
pp. 4081-4090 ◽  
Author(s):  
Shinya Nishi ◽  
Sheau Yu Hsu ◽  
Karen Zell ◽  
Aaron J. W. Hsueh
Keyword(s):  
G Protein ◽  
Hormone Receptors ◽  
Coupling Mechanism ◽  
Leucine Rich Repeat ◽  
G Protein Coupled Receptor ◽  
Glycoprotein Hormone ◽  
Glycoprotein Hormone Receptors ◽  
Signaling Mechanisms ◽  
G Protein Coupled

Abstract The receptors for lutropin (LH), FSH, and TSH belong to the large G protein-coupled receptor (GPCR) superfamily and are unique in having a large N-terminal extracellular (ecto-) domain important for interactions with the large glycoprotein hormone ligands. Recent studies indicated the evolution of a large family of the leucine-rich repeat-containing, G protein-coupled receptors (LGRs) with at least seven members in mammals. Based on the sequences of mammalian glycoprotein hormone receptors, we have identified a new LGR in Drosophila melanogaster and named it as fly LGR2 to distinguish it from the previously reported fly LH/FSH/TSH receptor (renamed as fly LGR1). Genomic analysis indicated the presence of 10 exons in fly LGR2 as compared with 16 exons in fly LGR1. The deduced fly LGR2 complementary DNA (cDNA) showed 43 and 64% similarity to the fly LGR1 in the ectodomain and transmembrane region, respectively. Comparison of 12 LGRs from diverse species indicated that these proteins can be divided into three subfamilies and fly LGR1 and LGR2 belong to different subfamilies. Potential signaling mechanisms were tested in human 293T cells overexpressing the fly receptors. Of interest, fly LGR1, but not LGR2, showed constitutive activity as reflected by elevated basal cAMP production in transfected cells. The basal activity of fly LGR1 was further augmented following point mutations of key residues in the intracellular loop 3 or transmembrane VI, similar to those found in patients with familial male precocious puberty. The present study reports the cloning of fly LGR2 and indicates that the G protein-coupling mechanism is conserved in fly LGR1 as compared with the mammalian glycoprotein hormone receptors. The characterization of fly receptors with features similar to mammalian glycoprotein hormone receptors allows a better understanding of the evolution of this unique group of GPCRs and future elucidation of their ligand signaling mechanisms.

Selective recruitment of arrestin-3 to clathrin coated pits upon stimulation of G protein-coupled receptors

2000 ◽  
Vol 113 (13) ◽  
pp. 2463-2470 ◽  
Author(s):  
F. Santini ◽  
R.B. Penn ◽  
A.W. Gagnon ◽  
J.L. Benovic ◽  
J.H. Keen
Keyword(s):  
G Protein ◽  
G Protein Coupled Receptors ◽  
Coated Pits ◽  
Over Expression ◽  
Physiological Conditions ◽  
A Cell ◽  
G Protein Coupled ◽  
Comparable Magnitude

Non-visual arrestins (arrestin-2 and arrestin-3) play critical roles in the desensitization and internalization of many G protein-coupled receptors. In vitro experiments have shown that both non-visual arrestins bind with high and approximately comparable affinities to activated, phosphorylated forms of receptors. They also exhibit high affinity binding, again of comparable magnitude, to clathrin. Further, agonist-promoted internalization of many receptors has been found to be stimulated by exogenous over-expression of either arrestin2 or arrestin3. The existence of multiple arrestins raises the question whether stimulated receptors are selective for a specific endogenous arrestin under more physiological conditions. Here we address this question in RBL-2H3 cells, a cell line that expresses comparable levels of endogenous arrestin-2 and arrestin-3. When (beta)(2)-adrenergic receptors are stably expressed in these cells the receptors internalize efficiently following agonist stimulation. However, by immunofluorescence microscopy we determine that only arrestin-3, but not arrestin-2, is rapidly recruited to clathrin coated pits upon receptor stimulation. Similarly, in RBL-2H3 cells that stably express physiological levels of m1AChR, the addition of carbachol selectively induces the localization of arrestin-3, but not arrestin-2, to coated pits. Thus, this work demonstrates coupling of G protein-coupled receptors to a specific non-visual arrestin in an in vivo setting.

N-ethyl-N-nitrosourea-based generation of mouse models for mutant G protein-coupled receptors

2006 ◽  
Vol 26 (3) ◽  
pp. 209-217 ◽  
Author(s):  
Johannes Grosse ◽  
Patrick Tarnow ◽  
Holger Römpler ◽  
Boris Schneider ◽  
Reinhard Sedlmeier ◽  
...  
Keyword(s):  
G Protein ◽  
Mouse Models ◽  
Germ Line ◽  
Random Mutagenesis ◽  
G Protein Coupled Receptors ◽  
In Vitro Assays ◽  
Vitro Fertilization ◽  
Type 3 ◽  
G Protein Coupled

Chemical random mutagenesis techniques with the germ line supermutagen N-ethyl- N-nitrosourea (ENU) have been established to provide comprehensive collections of mouse models, which were then mined and analyzed in phenotype-driven studies. Here, we applied ENU mutagenesis in a high-throughput fashion for a gene-driven identification of new mutations. Selected members of the large superfamily of G protein-coupled receptors (GPCR), melanocortin type 3 (Mc3r) and type 4 (Mc4r) receptors, and the orphan chemoattractant receptor GPR33, were used as model targets to prove the feasibility of this approach. Parallel archives of DNA and sperm from mice mutagenized with ENU were screened for mutations in these GPCR, and in vitro assays served as a preselection step before in vitro fertilization was performed to generate the appropriate mouse model. For example, mouse models for inherited obesity were established by selecting fully or partially inactivating mutations in Mc4r. Our technology described herein has the potential to provide mouse models for a GPCR dysfunction of choice within <4 mo and can be extended to other gene classes of interest.

scholarly journals Rationally designed chemokine-based toxin targeting the viral G protein-coupled receptor US28 potently inhibits cytomegalovirus infection in vivo

2015 ◽  
Vol 112 (27) ◽  
pp. 8427-8432 ◽  
Author(s):  
Katja Spiess ◽  
Mads G. Jeppesen ◽  
Mikkel Malmgaard-Clausen ◽  
Karen Krzywkowski ◽  
Kalpana Dulal ◽  
...  
Keyword(s):  
G Protein ◽  
Pseudomonas Exotoxin ◽  
Viral Pathogen ◽  
Virus Origin ◽  
Ligand Interactions ◽  
Toxin Protein ◽  
Novel Strategy ◽  
G Protein Coupled

The use of receptor–ligand interactions to direct toxins to kill diseased cells selectively has shown considerable promise for treatment of a number of cancers and, more recently, autoimmune disease. Here we move the fusion toxin protein (FTP) technology beyond cancer/autoimmune therapeutics to target the human viral pathogen, human cytomegalovirus (HCMV), on the basis of its expression of the 7TM G protein-coupled chemokine receptor US28. The virus origin of US28 provides an exceptional chemokine-binding profile with high selectivity and improved binding for the CX3C chemokine, CX3CL1. Moreover, US28 is constitutively internalizing by nature, providing highly effective FTP delivery. We designed a synthetic CX3CL1 variant engineered to have ultra-high affinity for US28 and greater specificity for US28 than the natural sole receptor for CX3CL1, CX3CR1, and we fused the synthetic variant with the cytotoxic domain of Pseudomonas Exotoxin A. This novel strategy of a rationally designed FTP provided unparalleled anti-HCMV efficacy and potency in vitro and in vivo.

Sign in / Sign up

Export Citation Format

Share Document