The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling

Ana Dorrego-Rivas; Jerome Ezan; Maïté M. Moreau; Sonia Poirault-Chassac; Nathalie Aubailly; Julie De Neve; Camille Blanchard; Francis Castets; Amélie Fréal; Arne Battefeld; Nathalie Sans; Mireille Montcouquiol

doi:10.1126/sciadv.abo6333

The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling

Ana Dorrego-Rivas, Jerome Ezan, Maïté M. Moreau, Sonia Poirault-Chassac, Nathalie Aubailly, Julie De Neve, Camille Blanchard, Francis Castets, Amélie Fréal, Arne Battefeld, Nathalie Sans, Mireille Montcouquiol^*

^*Corresponding author for this work

Research output: Contribution to Journal › Article › Academic › peer-review

Abstract

Core planar cell polarity (PCP) genes, which are involved in various neurodevelopmental disorders such as neural tube closure, epilepsy, and autism spectrum disorder, have poorly defined molecular signatures in neurons, mostly synapse-centric. Here, we show that the core PCP protein Prickle-like protein 2 (Prickle2) controls neuronal polarity and is a previously unidentified member of the axonal initial segment (AIS) proteome. We found that Prickle2 is present and colocalizes with AnkG480, the AIS master organizer, in the earliest stages of axonal specification and AIS formation. Furthermore, by binding to and regulating AnkG480, Prickle2 modulates its ability to bundle microtubules, a crucial mechanism for establishing neuronal polarity and AIS formation. Prickle2 depletion alters cytoskeleton organization, and Prickle2 levels determine both axon number and AIS maturation. Last, early Prickle2 depletion produces impaired action potential firing.

Original language	English
Article number	eabo6333
Pages (from-to)	1-21
Number of pages	21
Journal	Science advances
Volume	8
Issue number	36
DOIs	https://doi.org/10.1126/sciadv.abo6333
Publication status	Published - 9 Sept 2022

Bibliographical note

Funding Information:
We thank D. Wu (NIDCD/NIH, USA) for supplying Prickle2 antibody, V. Bennett (Duke Cancer Institute, USA) for supplying the AnkG270-GFP and AnkG190-GFP constructs, C. Hoogenraad (Utrecht University, The Netherlands) for the AnkG480-GFP cDNA, and M. Deans (University of Utah, USA) for GFP-Prickle2 cDNA construct. We thank C. Medina, A. Barranger, I. Seynat, S. Dos Santos Carvalho, J. Stanic, and C. Lefranc for technical assistance. We thank the personnel of the Animal Facility of the Neurocentre Magendie for the rat care. We thank the Bordeaux Neurocampus core facilities used in this study and supported by the LabEx BRAIN (grant ANR-10-LABX-43), including the Biochemistry facility BioProt, and the assistance of Y. Rufin for GST production and purification is acknowledged. We thank E. Pacary for initial advices of IUE in mice. Confocal and STORM imaging were done at the Bordeaux Imaging Center, a service unit of the CNRS-INSERM and Bordeaux University, member of the national infrastructure France BioImaging supported by the French National Research Agency (ANR-10-INBS-04). The help of F. Cordelières for the AIS macro for ImageJ and M. Mondin for STORM imaging is acknowledged. All of the above facilities were funded by the LabEx BRAIN (ANR-10-LABX-43). We thank M. Grubb and M. Engelhardt for supportive and constructive comments and C. Leterrier for advices on actin staining and imaging on STORM. This work was supported by Institut National de la Santé et de la Recherche Médicale INSERM (to M.M. and N.S.), Fondation pour la Recherche Médicale (FRM) “Equipe FRM 2016” DEQ20160334899 (to M.M.), Ministère de l’Enseignement Supérieur et de la Recherche (MESRI) (to A.D.-R.), LabEx BRAIN ANR-10-LABX-43 (to A.D.-R.), Bordeaux Neurocampus Startup grant by the Region Aquitaine 2018.599 (to A.B.), and IDEX ATTRACTIVITE Chaires Neurocampus of the University of Bordeaux (to A.B.). The Planar Polarity Team is part of the GPR Brain_2030 Nanocoding project.

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Dorrego-Rivas, A., Ezan, J., Moreau, M. M., Poirault-Chassac, S., Aubailly, N., De Neve, J., Blanchard, C., Castets, F., Fréal, A., Battefeld, A., Sans, N., & Montcouquiol, M. (2022). The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling. Science advances, 8(36), 1-21. Article eabo6333. https://doi.org/10.1126/sciadv.abo6333

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title = "The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling",

abstract = "Core planar cell polarity (PCP) genes, which are involved in various neurodevelopmental disorders such as neural tube closure, epilepsy, and autism spectrum disorder, have poorly defined molecular signatures in neurons, mostly synapse-centric. Here, we show that the core PCP protein Prickle-like protein 2 (Prickle2) controls neuronal polarity and is a previously unidentified member of the axonal initial segment (AIS) proteome. We found that Prickle2 is present and colocalizes with AnkG480, the AIS master organizer, in the earliest stages of axonal specification and AIS formation. Furthermore, by binding to and regulating AnkG480, Prickle2 modulates its ability to bundle microtubules, a crucial mechanism for establishing neuronal polarity and AIS formation. Prickle2 depletion alters cytoskeleton organization, and Prickle2 levels determine both axon number and AIS maturation. Last, early Prickle2 depletion produces impaired action potential firing.",

author = "Ana Dorrego-Rivas and Jerome Ezan and Moreau, {Ma{\"i}t{\'e} M.} and Sonia Poirault-Chassac and Nathalie Aubailly and {De Neve}, Julie and Camille Blanchard and Francis Castets and Am{\'e}lie Fr{\'e}al and Arne Battefeld and Nathalie Sans and Mireille Montcouquiol",

note = "Funding Information: We thank D. Wu (NIDCD/NIH, USA) for supplying Prickle2 antibody, V. Bennett (Duke Cancer Institute, USA) for supplying the AnkG270-GFP and AnkG190-GFP constructs, C. Hoogenraad (Utrecht University, The Netherlands) for the AnkG480-GFP cDNA, and M. Deans (University of Utah, USA) for GFP-Prickle2 cDNA construct. We thank C. Medina, A. Barranger, I. Seynat, S. Dos Santos Carvalho, J. Stanic, and C. Lefranc for technical assistance. We thank the personnel of the Animal Facility of the Neurocentre Magendie for the rat care. We thank the Bordeaux Neurocampus core facilities used in this study and supported by the LabEx BRAIN (grant ANR-10-LABX-43), including the Biochemistry facility BioProt, and the assistance of Y. Rufin for GST production and purification is acknowledged. We thank E. Pacary for initial advices of IUE in mice. Confocal and STORM imaging were done at the Bordeaux Imaging Center, a service unit of the CNRS-INSERM and Bordeaux University, member of the national infrastructure France BioImaging supported by the French National Research Agency (ANR-10-INBS-04). The help of F. Cordeli{\`e}res for the AIS macro for ImageJ and M. Mondin for STORM imaging is acknowledged. All of the above facilities were funded by the LabEx BRAIN (ANR-10-LABX-43). We thank M. Grubb and M. Engelhardt for supportive and constructive comments and C. Leterrier for advices on actin staining and imaging on STORM. This work was supported by Institut National de la Sant{\'e} et de la Recherche M{\'e}dicale INSERM (to M.M. and N.S.), Fondation pour la Recherche M{\'e}dicale (FRM) “Equipe FRM 2016” DEQ20160334899 (to M.M.), Minist{\`e}re de l{\textquoteright}Enseignement Sup{\'e}rieur et de la Recherche (MESRI) (to A.D.-R.), LabEx BRAIN ANR-10-LABX-43 (to A.D.-R.), Bordeaux Neurocampus Startup grant by the Region Aquitaine 2018.599 (to A.B.), and IDEX ATTRACTIVITE Chaires Neurocampus of the University of Bordeaux (to A.B.). The Planar Polarity Team is part of the GPR Brain_2030 Nanocoding project. Publisher Copyright: Copyright {\textcopyright} 2022 The Authors, some rights reserved.",

year = "2022",

month = sep,

day = "9",

doi = "10.1126/sciadv.abo6333",

language = "English",

volume = "8",

pages = "1--21",

journal = "Science advances",

issn = "2375-2548",

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Dorrego-Rivas, A, Ezan, J, Moreau, MM, Poirault-Chassac, S, Aubailly, N, De Neve, J, Blanchard, C, Castets, F, Fréal, A, Battefeld, A, Sans, N & Montcouquiol, M 2022, 'The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling', Science advances, vol. 8, no. 36, eabo6333, pp. 1-21. https://doi.org/10.1126/sciadv.abo6333

TY - JOUR

T1 - The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling

AU - Dorrego-Rivas, Ana

AU - Ezan, Jerome

AU - Moreau, Maïté M.

AU - Poirault-Chassac, Sonia

AU - Aubailly, Nathalie

AU - De Neve, Julie

AU - Blanchard, Camille

AU - Castets, Francis

AU - Fréal, Amélie

AU - Battefeld, Arne

AU - Sans, Nathalie

AU - Montcouquiol, Mireille

N1 - Funding Information: We thank D. Wu (NIDCD/NIH, USA) for supplying Prickle2 antibody, V. Bennett (Duke Cancer Institute, USA) for supplying the AnkG270-GFP and AnkG190-GFP constructs, C. Hoogenraad (Utrecht University, The Netherlands) for the AnkG480-GFP cDNA, and M. Deans (University of Utah, USA) for GFP-Prickle2 cDNA construct. We thank C. Medina, A. Barranger, I. Seynat, S. Dos Santos Carvalho, J. Stanic, and C. Lefranc for technical assistance. We thank the personnel of the Animal Facility of the Neurocentre Magendie for the rat care. We thank the Bordeaux Neurocampus core facilities used in this study and supported by the LabEx BRAIN (grant ANR-10-LABX-43), including the Biochemistry facility BioProt, and the assistance of Y. Rufin for GST production and purification is acknowledged. We thank E. Pacary for initial advices of IUE in mice. Confocal and STORM imaging were done at the Bordeaux Imaging Center, a service unit of the CNRS-INSERM and Bordeaux University, member of the national infrastructure France BioImaging supported by the French National Research Agency (ANR-10-INBS-04). The help of F. Cordelières for the AIS macro for ImageJ and M. Mondin for STORM imaging is acknowledged. All of the above facilities were funded by the LabEx BRAIN (ANR-10-LABX-43). We thank M. Grubb and M. Engelhardt for supportive and constructive comments and C. Leterrier for advices on actin staining and imaging on STORM. This work was supported by Institut National de la Santé et de la Recherche Médicale INSERM (to M.M. and N.S.), Fondation pour la Recherche Médicale (FRM) “Equipe FRM 2016” DEQ20160334899 (to M.M.), Ministère de l’Enseignement Supérieur et de la Recherche (MESRI) (to A.D.-R.), LabEx BRAIN ANR-10-LABX-43 (to A.D.-R.), Bordeaux Neurocampus Startup grant by the Region Aquitaine 2018.599 (to A.B.), and IDEX ATTRACTIVITE Chaires Neurocampus of the University of Bordeaux (to A.B.). The Planar Polarity Team is part of the GPR Brain_2030 Nanocoding project. Publisher Copyright: Copyright © 2022 The Authors, some rights reserved.

PY - 2022/9/9

Y1 - 2022/9/9

N2 - Core planar cell polarity (PCP) genes, which are involved in various neurodevelopmental disorders such as neural tube closure, epilepsy, and autism spectrum disorder, have poorly defined molecular signatures in neurons, mostly synapse-centric. Here, we show that the core PCP protein Prickle-like protein 2 (Prickle2) controls neuronal polarity and is a previously unidentified member of the axonal initial segment (AIS) proteome. We found that Prickle2 is present and colocalizes with AnkG480, the AIS master organizer, in the earliest stages of axonal specification and AIS formation. Furthermore, by binding to and regulating AnkG480, Prickle2 modulates its ability to bundle microtubules, a crucial mechanism for establishing neuronal polarity and AIS formation. Prickle2 depletion alters cytoskeleton organization, and Prickle2 levels determine both axon number and AIS maturation. Last, early Prickle2 depletion produces impaired action potential firing.

AB - Core planar cell polarity (PCP) genes, which are involved in various neurodevelopmental disorders such as neural tube closure, epilepsy, and autism spectrum disorder, have poorly defined molecular signatures in neurons, mostly synapse-centric. Here, we show that the core PCP protein Prickle-like protein 2 (Prickle2) controls neuronal polarity and is a previously unidentified member of the axonal initial segment (AIS) proteome. We found that Prickle2 is present and colocalizes with AnkG480, the AIS master organizer, in the earliest stages of axonal specification and AIS formation. Furthermore, by binding to and regulating AnkG480, Prickle2 modulates its ability to bundle microtubules, a crucial mechanism for establishing neuronal polarity and AIS formation. Prickle2 depletion alters cytoskeleton organization, and Prickle2 levels determine both axon number and AIS maturation. Last, early Prickle2 depletion produces impaired action potential firing.

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The core PCP protein Prickle2 regulates axon number and AIS maturation by binding to AnkG and modulating microtubule bundling

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