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1.
Methods Mol Biol ; 1947: 257-267, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30969421

RESUMO

Intracellular signal transduced by G protein-coupled receptors (GPCRs) is tightly controlled by a guanine nucleotide-binding complex made of G protein Gα, Gß, and Gγ subunits, as well as a growing array of regulatory and accessory proteins such as arrestins. G protein-independent ß-arrestin recruitment at GPCRs is universally accepted as the canonical interactor system and it has been found to be a powerful tracker of most GPCRs activation. Pharmacological concepts have evolved remarkably after the finding that different ligands, binding at the same receptor, can selectively activate specific subsets of signaling pathways among all pathways activated by balanced ligands. This new paradigm referred to as functional selectivity or biased signaling, has opened new avenues for the design of tailored drugs with enhanced therapeutic efficacies and reduced side effects. Here, we describe a unique platform for the interrogation of GPCR using a transcriptional-based assay to measure transient ß-arrestin recruitment called Tango.


Assuntos
Bioensaio/métodos , Receptores Acoplados a Proteínas-G/metabolismo , beta-Arrestinas/metabolismo , Bleomicina/farmacologia , Células HEK293 , Humanos , Ligantes , Transdução de Sinais , Ativação Transcricional
2.
Methods Mol Biol ; 1947: 323-336, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30969425

RESUMO

G protein-coupled receptors (GPCRs) comprise the largest family of integral membrane proteins, which are coupled to heterotrimeric G proteins to influence cell signaling. Subsequent to G protein activation, agonist-stimulated G protein-coupled receptor kinase (GRK) phosphorylation results in the recruitment of ß-arrestin proteins, which form both stable and unstable complexes with GPCRs. ß-Arrestins when bound to GPCRs not only contribute to the uncoupling of G protein signaling but also to the redistribution of GPCRs to clathrin-coated pits via their association with both clathrin and ß2-adaptin facilitating GPCR endocytosis. This allows ß-arrestins to couple GPCRs to additional cell signaling proteins allowing a second wave of receptor signaling. Importantly, the ß-arrestin-regulated subcellular localization of these complexes also plays a critical role in regulating how these signals are transduced and which proteins are recruited. Here, we describe a methodology for assessing the GPCR subcellular localization by super-resolution microscopy and suggest that this methodology can be extended to the study of GPCR/protein complexes.


Assuntos
Clatrina/metabolismo , Processamento de Imagem Assistida por Computador/métodos , Microscopia/métodos , Receptores Acoplados a Proteínas-G/metabolismo , beta-Arrestinas/metabolismo , Endocitose , Humanos , Transdução de Sinais
3.
Methods Mol Biol ; 1957: 3-8, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919344

RESUMO

Arrestins have now been implicated in the actions of virtually every G protein-coupled receptor (GPCR) for which they have been examined. Originally discovered for their role in the turnoff of visual perception, their newly discovered pleotropic functions in the cellular and physiological actions of GPCRs not only illuminate new mechanisms of signal transduction but also offer new avenues for therapeutic utility. Below, in this introductory chapter, we provide a short historical description and synopsis of how arrestins conceptually became associated with the function of GPCRs.


Assuntos
beta-Arrestinas/história , beta-Arrestinas/metabolismo , Animais , História do Século XX , Humanos , beta-Arrestinas/uso terapêutico
4.
Methods Mol Biol ; 1957: 69-82, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919347

RESUMO

Arrestin proteins were originally characterized as regulators of GPCR desensitization, and that function alone was sufficient to promote extreme interest in their study. It is now appreciated that arrestins also function as mediators of GPCR trafficking and G protein-independent signaling. This latter function places them as prominent players in the emerging field of qualitative signaling, which promises to launch a new area of pharmacology that defines ligands with selectivity/bias toward either G protein-dependent or -independent signaling. To meet the demands of research into arrestin function, methodology has evolved accordingly over the last three decades since the discovery of the arrestin family. Herein we describe state-of-the-art approaches for studying the role of arrestins (ß-arrestin1 aka arrestin 2, ß-arrestin2 aka arrestin 3) in GPCR function in a primary cell type, cultured airway smooth muscle cells.


Assuntos
Biologia Molecular/métodos , Músculo Liso/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Sistema Respiratório/metabolismo , beta-Arrestinas/metabolismo , Genes Dominantes , Humanos , RNA Interferente Pequeno/metabolismo , Retroviridae/metabolismo
5.
Methods Mol Biol ; 1957: 9-55, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919345

RESUMO

The ß-arrestins (ß-arrs) were initially appreciated for the roles they play in the desensitization and endocytosis of G protein-coupled receptors (GPCRs). They are now also known to act as multifunctional adaptor proteins binding many non-receptor protein partners to control multiple signalling pathways. ß-arrs therefore act as key regulatory hubs at the crossroads of external cell inputs and functional outputs in cellular processes ranging from gene transcription to cell growth, survival, cytoskeletal regulation, polarity, and migration. An increasing number of studies have also highlighted the scaffolding roles ß-arrs play in vivo in both physiological and pathological conditions, which opens up therapeutic avenues to explore. In this introductory review chapter, we discuss the functional roles that ß-arrs exert to control GPCR function, their dynamic scaffolding roles and how this impacts signal transduction events, compartmentalization of ß-arrs, how ß-arrs are regulated themselves, and how the combination of these events culminates in cellular regulation.


Assuntos
Células/metabolismo , Transdução de Sinais , beta-Arrestinas/metabolismo , Animais , Endocitose , Humanos , Modelos Biológicos , Transporte Proteico , beta-Arrestinas/química
6.
Methods Mol Biol ; 1957: 59-68, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919346

RESUMO

ß-Arrestins are adaptors that regulate the signaling and trafficking of G protein-coupled receptors (GPCRs). Bioluminescence resonance energy transfer (BRET) is a sensitive and versatile method for real-time monitoring of protein-protein interactions and protein kinesis within live cells, such as the recruitment of ß-arrestins to activated receptors at the plasma membrane (PM) and the trafficking of GPCR/ß-arrestin complexes to endosomes. Trafficking of receptor/ß-arrestin complexes can be assessed by BRET through tagging ß-arrestins with the donor luciferase from Renilla reniformis (Rluc) and anchoring the acceptor green fluorescent protein from the same species (rGFP) in distinct cell compartments (e.g., PM or endosomes) to generate highly efficient bystander BRET (referred to as enhanced bystander BRET (EbBRET)) upon re-localization of ß-arrestins to these compartments following receptor activation. Here, we outline the protocol for quantitatively monitoring ß-arrestin recruitment to agonist-activated Angiotensin II type 1 receptor (AT1R) and ß2-adrenergic receptor (ß2AR) at the PM and the trafficking of receptor/ß-arrestin complexes into endosomes using EbBRET-based biosensors.


Assuntos
Técnicas de Transferência de Energia por Ressonância de Bioluminescência/métodos , Receptores Acoplados a Proteínas-G/metabolismo , beta-Arrestinas/metabolismo , Membrana Celular/metabolismo , Células HEK293 , Humanos , Cinética , Ligantes , Transporte Proteico , Fatores de Tempo , beta-Arrestinas/agonistas
7.
Methods Mol Biol ; 1957: 83-91, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919348

RESUMO

Initially identified as monomers, G protein-coupled receptors (GPCRs) can also form functional dimers that act as distinct signalling hubs for the integration of cellular signalling. We previously found that the angiotensin II (Ang II) type 1 receptor (AT1R) and the prostaglandin F2α (PGF2α) receptor (FP), both important in the control of smooth muscle contractility, form such a functional heterodimeric complex in HEK 293 and vascular smooth muscle cells (Goupil et al., J Biol Chem 290:3137-3148, 2015; Sleno et al., J Biol Chem 292:12139-12152, 2017). In addition to canonical G protein coupling, GPCRs recruit and engage ß-arrestin-dependent pathways. Using BRET-based biosensors, we demonstrate how to assess recruitment of ß-arrestin-1 and -2 to AT1R and the AT1R/FP dimer in response to Ang II. Surprisingly, ß-arrestin-1 and -2 were recruited to the dimer, in response to PGF2α as well, even though FP alone cannot recruit either ß-arrestin-1 and -2.


Assuntos
Técnicas de Transferência de Energia por Ressonância de Bioluminescência/métodos , Multimerização Proteica , Receptores Acoplados a Proteínas-G/metabolismo , beta-Arrestinas/metabolismo , Angiotensina II/farmacologia , Dinoprosta/farmacologia , Células HEK293 , Humanos , Multimerização Proteica/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/metabolismo
8.
Methods Mol Biol ; 1957: 93-104, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919349

RESUMO

Ubiquitination of G protein-coupled receptors (GPCRs) is an important dynamic posttranslational modification that has been linked to the intracellular trafficking of internalized GPCRs to lysosomes. Ubiquitination of GPCRs is mediated by specific E3 ubiquitin ligases that are scaffolded by the adaptor proteins called ß-arrestins. Traditionally, detection of GPCR ubiquitination is achieved by using ubiquitin antibodies to Western blot immunoprecipitates of detergent-solubilized GPCRs expressed in heterologous cells. However, studies have also shown that bioluminescence resonance energy transfer (BRET)-based techniques can reveal ubiquitination of GPCRs in intact cells and in real time. This chapter describes a step-by-step protocol to evaluate ubiquitination of GPCRs using the BRET methodology.


Assuntos
Técnicas de Transferência de Energia por Ressonância de Bioluminescência/métodos , Receptores Acoplados a Proteínas-G/metabolismo , Ubiquitinação , beta-Arrestinas/metabolismo , Análise de Dados , Células HEK293 , Humanos , Ligação Proteica , Ubiquitina/metabolismo
9.
Methods Mol Biol ; 1957: 139-158, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919352

RESUMO

ß-Arrestins 1 and 2 (ß-arr1 and ß-arr2) are ubiquitous proteins with common and distinct functions. They were initially identified as proteins recruited to stimulated G protein-coupled receptors (GPCRs), regulating their desensitization and internalization. The discovery that ß-arrs could also interact with more than 400 non-GPCR protein partners brought to light their central roles as multifunctional scaffold proteins regulating multiple signalling pathways from the plasma membrane to the nucleus, downstream of GPCRs or independently from these receptors. Through the regulation of the activities and subcellular localization of their binding partners, ß-arrs control various cell processes such as proliferation, cytoskeletal rearrangement, cell motility, and apoptosis. Thus, the identification of ß-arrs binding partners and the characterization of their mode of interaction in cells are central to the understanding of their function. Here we provide methods to explore the molecular interaction of ß-arrs with other proteins in cellulo.


Assuntos
Mapeamento de Interação de Proteínas/métodos , beta-Arrestinas/metabolismo , Técnicas de Transferência de Energia por Ressonância de Bioluminescência , Células HEK293 , Humanos , Imunoprecipitação , Ligação Proteica , Saccharomyces cerevisiae/metabolismo , Técnicas do Sistema de Duplo-Híbrido
10.
Methods Mol Biol ; 1957: 159-168, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919353

RESUMO

ADP-ribosylation factors (ARF) are GTPases that act to control the activation of numerous signaling events and cellular responses. The ARF6 isoform, present at the plasma membrane, can be activated by the angiotensin II type 1 receptor (AT1R), a process dependent upon ß-arrestin recruitment to the activated receptor. Here, we describe classical methods used to assess ß-arrestin-dependent activation of ARF6 following agonist stimulation of cells. In addition, because ARF6 and ß-arrestin can form a complex, we describe the procedures used to detect the interaction of ß-arrestin with this GTPase.


Assuntos
Fatores de Ribosilação do ADP/metabolismo , Citoesqueleto de Actina/metabolismo , Biologia Molecular/métodos , beta-Arrestinas/metabolismo , DNA Complementar/metabolismo , Ativação Enzimática , GTP Fosfo-Hidrolases/metabolismo , Guanosina Trifosfato/metabolismo , Células HEK293 , Humanos , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/metabolismo , Transporte Proteico , RNA Interferente Pequeno/metabolismo , Proteínas Recombinantes de Fusão/metabolismo
11.
Methods Mol Biol ; 1957: 169-175, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919354

RESUMO

G protein-coupled receptors (GPCRs) comprise the largest family of integral membrane proteins, which in addition to signaling via heterotrimeric G proteins can activate small G proteins both directly and indirectly. The activation of a variety of GPCRs leads to the translocation of Ral GDP dissociation stimulator (RalGDS) to the plasma membrane, where it functions as a guanine nucleotide exchange factor of RalA to promote membrane blebbing. The translocation of RalGDS is ß-arrestin-dependent and can be inhibited by either the expression of the ß-arrestin1 amino-terminal domain or the expression of RalGDS clone 284 (amino acid residues 616-768 of RalGDS). We describe here a methodology for assessing GPCR-dependent stimulation of RalGDS plasma membrane translocation.


Assuntos
Membrana Celular/metabolismo , Biologia Molecular/métodos , Receptores Acoplados a Proteínas-G/metabolismo , beta-Arrestinas/metabolismo , Fator ral de Troca do Nucleotídeo Guanina/metabolismo , Análise de Dados , Células HEK293 , Humanos , Receptor Tipo 1 de Angiotensina/metabolismo
12.
Methods Mol Biol ; 1957: 177-194, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919355

RESUMO

ß-arrestins are so-called hub proteins: they make complexes with many different partners, assembling functional complexes, and thereby fulfilling their biological function. The importance of this process in G protein-coupled receptor (GPCR) signalling has been fully demonstrated for many different receptors. For direct interactions, determining the interface regions, on ß-arrestins and on the partners, is crucial for understanding the function of the complex. Indeed, this brings information on which proteins can interact simultaneously with ß-arrestins, or, on the contrary, which partners are exclusive. We present here a method in two steps: protein-protein docking allows finding a limited number of peptides predicted to be involved in the interaction, and then experimental approaches that might be used for validating the prediction.


Assuntos
Biologia Molecular/métodos , beta-Arrestinas/metabolismo , Sequência de Aminoácidos , Fluorescência , Humanos , Interferometria , Peptídeos/química , Peptídeos/metabolismo , Ligação Proteica , Reprodutibilidade dos Testes , beta-Arrestinas/química
13.
Methods Mol Biol ; 1957: 195-215, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919356

RESUMO

Dynamic models of signaling networks allow the formulation of hypotheses on the topology and kinetic rate laws characterizing a given molecular network, in-depth exploration, and confrontation with kinetic biological data. Despite its standardization, dynamic modeling of signaling networks still requires successive technical steps that need to be carefully performed. Here, we detail these steps by going through the mathematical and statistical framework. We explain how it can be applied to the understanding of ß-arrestin-dependent signaling networks. We illustrate our methodology through the modeling of ß-arrestin recruitment kinetics at the follicle-stimulating hormone (FSH) receptor supported by in-house bioluminescence resonance energy transfer (BRET) data.


Assuntos
Modelos Biológicos , Transdução de Sinais , beta-Arrestinas/metabolismo , Técnicas de Transferência de Energia por Ressonância de Bioluminescência , Simulação por Computador , Células HEK293 , Humanos , Modelos Estatísticos
14.
Methods Mol Biol ; 1957: 217-232, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919357

RESUMO

Protein-protein interaction is crucial to protein function and cellular signaling. A number of approaches were developed and applied to characterize protein-protein interactions in the past decades. In recent years, mass spectrometry (MS)-based proteomic methods have emerged as powerful tools to identify protein binding partners in a global and high-throughput manner. In this chapter, we describe the proteomic methods used to characterize the whole sets of proteins associated with ß-arrestins (ß-arrestin interactomes). The method starts with co-immunoprecipitation (co-IP) of ß-arrestin signaling complexes from cells followed by protease digestion and LC/MS/MS analysis (liquid chromatography/tandem mass spectrometry) of proteins in the ß-arrestin signaling complexes. To investigate changes in the amounts of binding partners under different conditions, we also describe a SILAC (stable isotope labeling by amino acids in cell culture) method to obtain quantitative information for ß-arrestin interactomes.


Assuntos
Mapas de Interação de Proteínas , Proteômica/métodos , beta-Arrestinas/metabolismo , Biologia Computacional , Bases de Dados de Proteínas , Células HEK293 , Humanos , Transdução de Sinais , Tripsina/metabolismo
15.
Methods Mol Biol ; 1957: 251-269, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919359

RESUMO

ß-Arrestins (ß-arrs) were originally appreciated for the roles they play in the desensitization and internalization of G protein-coupled receptors (GPCRs). They are also now known to act as molecular scaffolds, providing control in multiple signalling pathways. Through their scaffolding properties, ß-arrs dynamically regulate the activity and/or subcellular distribution of protein partners giving rise to an appropriate cellular response. There are two ß-arr isoforms, namely, ß-arr1 and ß-arr2, which share high sequence homology and structural conservation. While the ß-arrs often display conserved overlapping roles, decisive differences between the isoforms also exist. A striking example of this is the subcellular distribution of the ß-arr isoforms. While ß-arr1 is distributed both in cytoplasmic and nuclear compartments, ß-arr2 displays an apparent cytoplasmic distribution. Both ß-arrs are actively imported into the nucleus, but ß-arr2 is constitutively exported by a leptomycin B-sensitive pathway due to a nuclear export signal in its C-terminus that is absent in ß-arr1. ß-arr2 therefore undergoes constitutive nucleocytoplasmic shuttling enabling the displacement of nuclear binding cargoes, such as Mdm2. Here, we describe methods to explore the differential nucleocytoplasmic shuttling capacities of the ß-arrs.


Assuntos
Núcleo Celular/metabolismo , Biologia Molecular/métodos , beta-Arrestinas/metabolismo , Transporte Ativo do Núcleo Celular , Sequência de Aminoácidos , Bioensaio , Imunofluorescência , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Células HeLa , Humanos , Processamento de Imagem Assistida por Computador , Cinética , Modelos Biológicos , Mutagênese/genética , Saccharomyces cerevisiae/metabolismo , Transdução de Sinais , Transformação Genética , beta-Arrestinas/química
16.
Methods Mol Biol ; 1957: 293-308, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919361

RESUMO

ß-Arrestins are key regulation proteins for G protein-coupled receptors (GPCRs) signaling. Experimental evidence suggests that ß-arrestins undergo conformational changes concomitant with binding to activated, phosphorylated GPCRs. We developed a mass spectrometry-based structural proteomic assay to monitor conformational changes associated with the activation of ß-arrestins. This assay utilizes synthesized phosphopeptides mimicking phosphorylated C-terminal tails of GPCRs to activate ß-arrestins. The activation-dependent conformational changes of ß-arrestins are revealed using limited proteolysis coupled with both SDS-PAGE and mass spectrometry analysis. As an in vitro ß-arrestin activation assay, this mass spectrometry-based structural method can be adapted as a simple but useful tool to study the nature and extent of conformational changes of ß-arrestins downstream of different receptors as well as ß-arrestin conformations associated with different functions, such as desensitization, internalization, and signaling.


Assuntos
Bioensaio/métodos , Espectrometria de Massas/métodos , beta-Arrestinas/química , Sequência de Aminoácidos , Análise de Dados , Fosfopeptídeos/síntese química , Fosfopeptídeos/química , Conformação Proteica , Proteólise , Tripsina/metabolismo , beta-Arrestinas/metabolismo
17.
Methods Mol Biol ; 1957: 325-334, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919363

RESUMO

Neisseria meningitidis is a Gram-negative diplococcus restricted to humans that causes severe septicemia and/or meningitidis. Initial adhesion to human endothelial cells is mediated through the interaction of type IV pili with the hetero-oligomeric complexes formed by the human receptors CD147 and the ß2-adrenergic receptor. Interaction with this complex heterodimer activates a ß-arrestin-biased signaling pathway leading to actin polymerization and accumulation of ezrin and ezrin-binding partners. These signaling events promote the formation of cell plasma membrane protrusions in endothelial cells, which are crucial for N. meningitidis colonies to resist shear stress and colonize blood vessels. Here we provide detailed protocols to evaluate the role of ß-arrestins in actin and ezrin signaling downstream of G protein-coupled receptor activation.


Assuntos
Biologia Molecular/métodos , Neisseria meningitidis/metabolismo , Transdução de Sinais , beta-Arrestinas/metabolismo , Células Cultivadas , Células Endoteliais/metabolismo , Células Endoteliais/microbiologia , Células HEK293 , Humanos , RNA Interferente Pequeno/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo
18.
Methods Mol Biol ; 1957: 345-364, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919365

RESUMO

Novel findings reveal important functional roles for ß-arrestin 1 and ß-arrestin 2 in the regulation of insulin secretion, ß-cell survival, and ß-cell mass plasticity not only by glucose but also by G-protein-coupled receptors, such as the glucagon-like peptide-1 (GLP-1) and the pituitary adenylate cyclase-activating polypeptide (PACAP) receptors or GPR40, or tyrosine kinase receptors, such as the insulin receptor. Here, we describe experimental protocols to knock down ß-arrestins by small interference RNA, to follow subcellular localization of ß-arrestins in the cytosol and nucleus of the insulinoma INS-1E rat pancreatic ß-cell line, and to analyze ß-arrestin protein expression by Western blot using INS-1E cells and isolated mouse or human pancreatic islets. We also provide details on how to genotype ß-arrestin 2 knockout (Arrb2-/-) mice and to evaluate ß-arrestin-mediated roles in ß-cell mass plasticity and ß-cell signaling using immunocytochemistry on pancreatic sections or on primary dispersed ß-cells from wild-type mice and Arrb2-/- mice.


Assuntos
Células Secretoras de Insulina/metabolismo , Biologia Molecular/métodos , beta-Arrestinas/metabolismo , Animais , Citoplasma/metabolismo , Técnicas de Silenciamento de Genes , Genótipo , Camundongos Knockout , Proteínas Nucleares/metabolismo , RNA Interferente Pequeno/metabolismo
19.
Methods Mol Biol ; 1957: 365-384, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919366

RESUMO

Type II diabetes is one of the most serious worldwide public health problems, and its hallmark is insulin resistance, obesity associated with chronic inflammation, and defective islet ß-cell function. ß-Arrestins play important roles in diabetes pathogenesis through scaffolding insulin-induced AKT activation in the liver, suppressing peroxisome proliferator-activated receptor-γ-mediated adipogenesis and inflammatory responses in adipose tissue and through promoting GLP-1-induced insulin secretion in the islet. The current chapter provides detailed protocols for both in vitro and in vivo studies of the function of ß-arrestins associated with type II diabetes.


Assuntos
Biologia Molecular/métodos , beta-Arrestinas/metabolismo , Adipócitos/metabolismo , Animais , Células da Medula Óssea/citologia , Células Cultivadas , Regulação da Expressão Gênica , Glucose/biossíntese , Hepatócitos/metabolismo , Resistência à Insulina , Secreção de Insulina , Células Secretoras de Insulina/metabolismo , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais , Coloração e Rotulagem
20.
Methods Mol Biol ; 1957: 385-391, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30919367

RESUMO

ß-Arrestin function has largely been investigated in cell culture-based systems. Here we describe methods to investigate ß-arrestin2 signaling in vivo by developing novel mouse models and viral methods to overexpress ß-arrestin2 in the mouse brain. The methods and concepts described here are in the context of Parkinson's disease (PD) and developing automated in vivo drug screening platforms.


Assuntos
Biologia Molecular/métodos , Doença de Parkinson/metabolismo , Transdução de Sinais , beta-Arrestinas/metabolismo , Animais , Modelos Animais de Doenças , Dopamina/deficiência , Discinesias/patologia , Discinesias/fisiopatologia , Humanos , Levodopa/farmacologia , Locomoção , Camundongos Knockout , Doença de Parkinson/fisiopatologia , Técnicas Estereotáxicas
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