A proteomic analysis of LRRK2 binding partners reveals interactions with multiple signaling components of the WNT-PCP pathwayReportar como inadecuado




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Molecular Neurodegeneration

, 12:54

First Online: 11 July 2017Received: 25 October 2016Accepted: 20 June 2017

Abstract

BackgroundAutosomal-dominant mutations in the Park8 gene encoding Leucine-rich repeat kinase 2 LRRK2 have been identified to cause up to 40% of the genetic forms of Parkinson’s disease. However, the function and molecular pathways regulated by LRRK2 are largely unknown. It has been shown that LRRK2 serves as a scaffold during activation of WNT-β-catenin signaling via its interaction with the β-catenin destruction complex, DVL1-3 and LRP6. In this study, we examine whether LRRK2 also interacts with signaling components of the WNT-Planar Cell Polarity WNT-PCP pathway, which controls the maturation of substantia nigra dopaminergic neurons, the main cell type lost in Parkinson’s disease patients.

MethodsCo-immunoprecipitation and tandem mass spectrometry was performed in a mouse substantia nigra cell line SN4741 and human HEK293T cell line in order to identify novel LRRK2 binding partners. Inhibition of the WNT-β-catenin reporter, TOPFlash, was used as a read-out of WNT-PCP pathway activation. The capacity of LRRK2 to regulate WNT-PCP signaling in vivo was tested in Xenopus laevis’ early development.

ResultsOur proteomic analysis identified that LRRK2 interacts with proteins involved in WNT-PCP signaling such as the PDZ domain-containing protein GIPC1 and Integrin-linked kinase ILK in dopaminergic cells in vitro and in the mouse ventral midbrain in vivo. Moreover, co-immunoprecipitation analysis revealed that LRRK2 binds to two core components of the WNT-PCP signaling pathway, PRICKLE1 and CELSR1, as well as to FLOTILLIN-2 and CULLIN-3, which regulate WNT secretion and inhibit WNT-β-catenin signaling, respectively. We also found that PRICKLE1 and LRRK2 localize in signalosomes and act as dual regulators of WNT-PCP and β-catenin signaling. Accordingly, analysis of the function of LRRK2 in vivo, in X. laevis revelaed that LRKK2 not only inhibits WNT-β-catenin pathway, but induces a classical WNT-PCP phenotype in vivo.

ConclusionsOur study shows for the first time that LRRK2 activates the WNT-PCP signaling pathway through its interaction to multiple WNT-PCP components. We suggest that LRRK2 regulates the balance between WNT-β-catenin and WNT-PCP signaling, depending on the binding partners. Since this balance is crucial for homeostasis of midbrain dopaminergic neurons, we hypothesize that its alteration may contribute to the pathophysiology of Parkinson’s disease.

KeywordsWNT-planar cell polarity PRICKLE1 CELSR1 DVL Parkinson’s disease Dopaminergic neurons Substantia nigra Immunoprecipitation Endocytosis Signalosomes AbbreviationsBEND3BEN domain-containing protein 3

CNSCentral nervous system

CORC-terminal of COR domain

DVLSegment polarity protein dishevelled homolog DVL

ENA-VASPEnabled-Vasodilator proteins

GIPC1PDZ domain-containing protein GIPC1

GPCRsG protein coupled receptors

ILKIntegrin-linked kinase

LPPLipoma-preferred partner

LRP6Low-density lipoprotein receptor-related protein 6

LRRLeucine-rich repeats

LRRK2 KDLRRK2 knock down

LRRK2 WTLRRK2 wild type

LRRK2Leucine-rich repeat kinase 2

mDA neuronsmouse dopaminergic neurons

PCPPlanar Cell Polarity

PDParkinson’s disease

ROCRas of complex protein domain

SDOCHSodium deoxycholate buffer

SNpcSubstantia nigra pars compacta

VANGL2Vang-like protein 2

VMVentral midbrain

WNTWingless-Integration

Electronic supplementary materialThe online version of this article doi:10.1186-s13024-017-0193-9 contains supplementary material, which is available to authorized users.

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Autor: Alena Salašová - Chika Yokota - David Potěšil - Zbyněk Zdráhal - Vítězslav Bryja - Ernest Arenas

Fuente: https://link.springer.com/







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