CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity

Wenting Guo; Haibo Wang; Arun Kumar Tharkeshwar; Julien Couthouis; Elke Braems; Pegah Masrori; Evelien Van Schoor; Yannan Fan; Karan Ahuja; Matthieu Moisse; Maarten Jacquemyn; Rodrigo Furtado Madeiro da Costa; Madhavsai Gajjar; Sriram Balusu; Tine Tricot; Laura Fumagalli; Nicole Hersmus; Rekin's Janky; Francis Impens; Pieter Vanden Berghe; Ritchie Ho; Dietmar Rudolf Thal; Rik Vandenberghe; Muralidhar L. Hegde; Siddharthan Chandran; Bart De Strooper; Dirk Daelemans; Philip Van Damme; Ludo Van Den Bosch; Catherine Verfaillie

doi:10.1002/alz.12760

CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity

Wenting Guo, Haibo Wang, Arun Kumar Tharkeshwar, Julien Couthouis, Elke Braems, Pegah Masrori, Evelien Van Schoor, Yannan Fan, Karan Ahuja, Matthieu Moisse, Maarten Jacquemyn, Rodrigo Furtado Madeiro da Costa, Madhavsai Gajjar, Sriram Balusu, Tine Tricot, Laura Fumagalli, Nicole Hersmus, Rekin's Janky, Francis Impens, Pieter Vanden BergheRitchie Ho, Dietmar Rudolf Thal, Rik Vandenberghe, Muralidhar L. Hegde, Siddharthan Chandran, Bart De Strooper, Dirk Daelemans, Philip Van Damme, Ludo Van Den Bosch, Catherine Verfaillie

Research output: Contribution to journal › Article › peer-review

14 Scopus citations

Abstract

Introduction: The most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are hexanucleotide repeats in chromosome 9 open reading frame 72 (C9orf72). These repeats produce dipeptide repeat proteins with poly(PR) being the most toxic one. Methods: We performed a kinome-wide CRISPR/Cas9 knock-out screen in human induced pluripotent stem cell (iPSC) -derived cortical neurons to identify modifiers of poly(PR) toxicity, and validated the role of candidate modifiers using in vitro, in vivo, and ex-vivo studies. Results: Knock-down of NIMA-related kinase 6 (NEK6) prevented neuronal toxicity caused by poly(PR). Knock-down of nek6 also ameliorated the poly(PR)-induced axonopathy in zebrafish and NEK6 was aberrantly expressed in C9orf72 patients. Suppression of NEK6 expression and NEK6 activity inhibition rescued axonal transport defects in cortical neurons from C9orf72 patient iPSCs, at least partially by reversing p53-related DNA damage. Discussion: We identified NEK6, which regulates poly(PR)-mediated p53-related DNA damage, as a novel therapeutic target for C9orf72 FTD/ALS.

Original language	English (US)
Pages (from-to)	1245-1259
Number of pages	15
Journal	Alzheimer's and Dementia
Volume	19
Issue number	4
DOIs	https://doi.org/10.1002/alz.12760
State	Published - Apr 2023

Keywords

C9orf72
CRISPR/Cas9 screen
DNA damage
NEK6
PR toxicity
amyotrophic lateral sclerosis
frontotemporal dementia
human pluripotent stem cells
neurodegeneration
p53
DNA Repeat Expansion/genetics
C9orf72 Protein/genetics
Humans
Tumor Suppressor Protein p53/genetics
Amyotrophic Lateral Sclerosis/genetics
Zebrafish/genetics
Frontotemporal Dementia/genetics
NIMA-Related Kinases/genetics
Animals
Neurons/metabolism
CRISPR-Cas Systems
Induced Pluripotent Stem Cells/metabolism

ASJC Scopus subject areas

Clinical Neurology
Geriatrics and Gerontology
Psychiatry and Mental health
Cellular and Molecular Neuroscience
Health Policy
Developmental Neuroscience
Epidemiology

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10.1002/alz.12760

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Guo, W., Wang, H., Kumar Tharkeshwar, A., Couthouis, J., Braems, E., Masrori, P., Van Schoor, E., Fan, Y., Ahuja, K., Moisse, M., Jacquemyn, M., Furtado Madeiro da Costa, R., Gajjar, M., Balusu, S., Tricot, T., Fumagalli, L., Hersmus, N., Janky, R., Impens, F., ... Verfaillie, C. (2023). CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity. Alzheimer's and Dementia, 19(4), 1245-1259. https://doi.org/10.1002/alz.12760

Guo, W, Wang, H, Kumar Tharkeshwar, A, Couthouis, J, Braems, E, Masrori, P, Van Schoor, E, Fan, Y, Ahuja, K, Moisse, M, Jacquemyn, M, Furtado Madeiro da Costa, R, Gajjar, M, Balusu, S, Tricot, T, Fumagalli, L, Hersmus, N, Janky, R, Impens, F, Vanden Berghe, P, Ho, R, Thal, DR, Vandenberghe, R, Hegde, ML, Chandran, S, De Strooper, B, Daelemans, D, Van Damme, P, Van Den Bosch, L & Verfaillie, C 2023, 'CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity', Alzheimer's and Dementia, vol. 19, no. 4, pp. 1245-1259. https://doi.org/10.1002/alz.12760

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title = "CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity",

abstract = "Introduction: The most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are hexanucleotide repeats in chromosome 9 open reading frame 72 (C9orf72). These repeats produce dipeptide repeat proteins with poly(PR) being the most toxic one. Methods: We performed a kinome-wide CRISPR/Cas9 knock-out screen in human induced pluripotent stem cell (iPSC) -derived cortical neurons to identify modifiers of poly(PR) toxicity, and validated the role of candidate modifiers using in vitro, in vivo, and ex-vivo studies. Results: Knock-down of NIMA-related kinase 6 (NEK6) prevented neuronal toxicity caused by poly(PR). Knock-down of nek6 also ameliorated the poly(PR)-induced axonopathy in zebrafish and NEK6 was aberrantly expressed in C9orf72 patients. Suppression of NEK6 expression and NEK6 activity inhibition rescued axonal transport defects in cortical neurons from C9orf72 patient iPSCs, at least partially by reversing p53-related DNA damage. Discussion: We identified NEK6, which regulates poly(PR)-mediated p53-related DNA damage, as a novel therapeutic target for C9orf72 FTD/ALS.",

keywords = "C9orf72, CRISPR/Cas9 screen, DNA damage, NEK6, PR toxicity, amyotrophic lateral sclerosis, frontotemporal dementia, human pluripotent stem cells, neurodegeneration, p53, DNA Repeat Expansion/genetics, C9orf72 Protein/genetics, Humans, Tumor Suppressor Protein p53/genetics, Amyotrophic Lateral Sclerosis/genetics, Zebrafish/genetics, Frontotemporal Dementia/genetics, NIMA-Related Kinases/genetics, Animals, Neurons/metabolism, CRISPR-Cas Systems, Induced Pluripotent Stem Cells/metabolism",

author = "Wenting Guo and Haibo Wang and {Kumar Tharkeshwar}, Arun and Julien Couthouis and Elke Braems and Pegah Masrori and {Van Schoor}, Evelien and Yannan Fan and Karan Ahuja and Matthieu Moisse and Maarten Jacquemyn and {Furtado Madeiro da Costa}, Rodrigo and Madhavsai Gajjar and Sriram Balusu and Tine Tricot and Laura Fumagalli and Nicole Hersmus and Rekin's Janky and Francis Impens and {Vanden Berghe}, Pieter and Ritchie Ho and Thal, {Dietmar Rudolf} and Rik Vandenberghe and Hegde, {Muralidhar L.} and Siddharthan Chandran and {De Strooper}, Bart and Dirk Daelemans and {Van Damme}, Philip and {Van Den Bosch}, Ludo and Catherine Verfaillie",

note = "{\textcopyright} 2022 The Authors. Alzheimer's & Dementia published by Wiley Periodicals LLC on behalf of Alzheimer's Association.",

year = "2023",

month = apr,

doi = "10.1002/alz.12760",

language = "English (US)",

volume = "19",

pages = "1245--1259",

journal = "Alzheimer's and Dementia",

issn = "1552-5260",

publisher = "Elsevier",

number = "4",

}

TY - JOUR

T1 - CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity

AU - Guo, Wenting

AU - Wang, Haibo

AU - Kumar Tharkeshwar, Arun

AU - Couthouis, Julien

AU - Braems, Elke

AU - Masrori, Pegah

AU - Van Schoor, Evelien

AU - Fan, Yannan

AU - Ahuja, Karan

AU - Moisse, Matthieu

AU - Jacquemyn, Maarten

AU - Furtado Madeiro da Costa, Rodrigo

AU - Gajjar, Madhavsai

AU - Balusu, Sriram

AU - Tricot, Tine

AU - Fumagalli, Laura

AU - Hersmus, Nicole

AU - Janky, Rekin's

AU - Impens, Francis

AU - Vanden Berghe, Pieter

AU - Ho, Ritchie

AU - Thal, Dietmar Rudolf

AU - Vandenberghe, Rik

AU - Hegde, Muralidhar L.

AU - Chandran, Siddharthan

AU - De Strooper, Bart

AU - Daelemans, Dirk

AU - Van Damme, Philip

AU - Van Den Bosch, Ludo

AU - Verfaillie, Catherine

PY - 2023/4

Y1 - 2023/4

N2 - Introduction: The most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are hexanucleotide repeats in chromosome 9 open reading frame 72 (C9orf72). These repeats produce dipeptide repeat proteins with poly(PR) being the most toxic one. Methods: We performed a kinome-wide CRISPR/Cas9 knock-out screen in human induced pluripotent stem cell (iPSC) -derived cortical neurons to identify modifiers of poly(PR) toxicity, and validated the role of candidate modifiers using in vitro, in vivo, and ex-vivo studies. Results: Knock-down of NIMA-related kinase 6 (NEK6) prevented neuronal toxicity caused by poly(PR). Knock-down of nek6 also ameliorated the poly(PR)-induced axonopathy in zebrafish and NEK6 was aberrantly expressed in C9orf72 patients. Suppression of NEK6 expression and NEK6 activity inhibition rescued axonal transport defects in cortical neurons from C9orf72 patient iPSCs, at least partially by reversing p53-related DNA damage. Discussion: We identified NEK6, which regulates poly(PR)-mediated p53-related DNA damage, as a novel therapeutic target for C9orf72 FTD/ALS.

AB - Introduction: The most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are hexanucleotide repeats in chromosome 9 open reading frame 72 (C9orf72). These repeats produce dipeptide repeat proteins with poly(PR) being the most toxic one. Methods: We performed a kinome-wide CRISPR/Cas9 knock-out screen in human induced pluripotent stem cell (iPSC) -derived cortical neurons to identify modifiers of poly(PR) toxicity, and validated the role of candidate modifiers using in vitro, in vivo, and ex-vivo studies. Results: Knock-down of NIMA-related kinase 6 (NEK6) prevented neuronal toxicity caused by poly(PR). Knock-down of nek6 also ameliorated the poly(PR)-induced axonopathy in zebrafish and NEK6 was aberrantly expressed in C9orf72 patients. Suppression of NEK6 expression and NEK6 activity inhibition rescued axonal transport defects in cortical neurons from C9orf72 patient iPSCs, at least partially by reversing p53-related DNA damage. Discussion: We identified NEK6, which regulates poly(PR)-mediated p53-related DNA damage, as a novel therapeutic target for C9orf72 FTD/ALS.

KW - C9orf72

KW - CRISPR/Cas9 screen

KW - DNA damage

KW - NEK6

KW - PR toxicity

KW - amyotrophic lateral sclerosis

KW - frontotemporal dementia

KW - human pluripotent stem cells

KW - neurodegeneration

KW - p53

KW - DNA Repeat Expansion/genetics

KW - C9orf72 Protein/genetics

KW - Humans

KW - Tumor Suppressor Protein p53/genetics

KW - Amyotrophic Lateral Sclerosis/genetics

KW - Zebrafish/genetics

KW - Frontotemporal Dementia/genetics

KW - NIMA-Related Kinases/genetics

KW - Animals

KW - Neurons/metabolism

KW - CRISPR-Cas Systems

KW - Induced Pluripotent Stem Cells/metabolism

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U2 - 10.1002/alz.12760

DO - 10.1002/alz.12760

M3 - Article

C2 - 35993441

AN - SCOPUS:85136490463

SN - 1552-5260

VL - 19

SP - 1245

EP - 1259

JO - Alzheimer's and Dementia

JF - Alzheimer's and Dementia

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ER -

CRISPR/Cas9 screen in human iPSC-derived cortical neurons identifies NEK6 as a novel disease modifier of C9orf72 poly(PR) toxicity

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ASJC Scopus subject areas

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