Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus

Lina Odqvist; Zala Jevnikar; Rebecca Riise; Lisa Öberg; Magdalena Rhedin; Dag Leonard; Linda Yrlid; Sonya Jackson; Johan Mattsson; Sambit Nanda; Philip Cohen; Axel Knebel; Simon Arthur; Kristofer Thörn; Elisabet Svenungsson; Andreas Jönsen; Iva Gunnarsson; Karolina Tandre; Andrei Alexsson; Alf Kastbom; Solbritt Rantapää-Dahlqvist; Maija-Leena Eloranta; Ann-Christine Syvänen; Anders Bengtsson; Patrik Johansson; Johanna K. Sandling; Christopher Sjöwall; Lars Rönnblom; Barry Collins; Outi Vaarala

doi:10.1136/annrheumdis-2019-215434

Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus

Lina Odqvist, Zala Jevnikar, Rebecca Riise, Lisa Öberg, Magdalena Rhedin, Dag Leonard, Linda Yrlid, Sonya Jackson, Johan Mattsson, Sambit Nanda, Philip Cohen, Axel Knebel, Simon Arthur, Kristofer Thörn, Elisabet Svenungsson, Andreas Jönsen, Iva Gunnarsson, Karolina Tandre, Andrei Alexsson, Alf KastbomSolbritt Rantapää-Dahlqvist, Maija-Leena Eloranta, Ann-Christine Syvänen, Anders Bengtsson, Patrik Johansson, Johanna K. Sandling, Christopher Sjöwall, Lars Rönnblom, Barry Collins, Outi Vaarala (Lead / Corresponding author)

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Abstract

Objectives: Genetic variations in TNFAIP3 (A20) de-ubiquitinase (DUB) domain increase the risk of systemic lupus erythematosus (SLE) and rheumatoid arthritis. A20 is a negative regulator of NF-κB but the role of its DUB domain and related genetic variants remain unclear. We aimed to study the functional effects of A20 DUB-domain alterations in immune cells and understand its link to SLE pathogenesis.

Methods: CRISPR/Cas9 was used to generate human U937 monocytes with A20 DUB-inactivating C103A knock-in (KI) mutation. Whole genome RNA-sequencing was used to identify differentially expressed genes between WT and C103A KI cells. Functional studies were performed in A20 C103A U937 cells and in immune cells from A20 C103A mice and genotyped healthy individuals with A20 DUB polymorphism rs2230926. Neutrophil extracellular trap (NET) formation was addressed ex vivo in neutrophils from A20 C103A mice and SLE-patients with rs2230926.

Results: Genetic disruption of A20 DUB domain in human and murine myeloid cells did not give rise to enhanced NF-κB signalling. Instead, cells with C103A mutation or rs2230926 polymorphism presented an upregulated expression of PADI4, an enzyme regulating protein citrullination and NET formation, two key mechanisms in autoimmune pathology. A20 C103A cells exhibited enhanced protein citrullination and extracellular trap formation, which could be suppressed by selective PAD4 inhibition. Moreover, SLE-patients with rs2230926 showed increased NETs and increased frequency of autoantibodies to citrullinated epitopes.

Conclusions: We propose that genetic alterations disrupting the A20 DUB domain mediate increased susceptibility to SLE through the upregulation of PADI4 with resultant protein citrullination and extracellular trap formation.

Original language	English
Pages (from-to)	1363-1370
Number of pages	8
Journal	Annals of the Rheumatic Diseases
Volume	78
Issue number	10
Early online date	12 Jul 2019
DOIs	https://doi.org/10.1136/annrheumdis-2019-215434
Publication status	Published - 1 Oct 2019

Keywords

NET
PAD4
PADI4
peptidyl arginine deiminase
rs2230926

ASJC Scopus subject areas

Rheumatology
Immunology and Allergy
Immunology
General Biochemistry,Genetics and Molecular Biology

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1136/annrheumdis-2019-215434Licence: CC BY-NC

Final Published VersionFinal published version, 2.04 MBLicence: CC BY-NC

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Odqvist, L., Jevnikar, Z., Riise, R., Öberg, L., Rhedin, M., Leonard, D., Yrlid, L., Jackson, S., Mattsson, J., Nanda, S., Cohen, P., Knebel, A., Arthur, S., Thörn, K., Svenungsson, E., Jönsen, A., Gunnarsson, I., Tandre, K., Alexsson, A., ... Vaarala, O. (2019). Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus. Annals of the Rheumatic Diseases, 78(10), 1363-1370. https://doi.org/10.1136/annrheumdis-2019-215434

@article{82bb6e565bcd472babfbacb4cce28cd1,

title = "Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus",

abstract = "Objectives: Genetic variations in TNFAIP3 (A20) de-ubiquitinase (DUB) domain increase the risk of systemic lupus erythematosus (SLE) and rheumatoid arthritis. A20 is a negative regulator of NF-κB but the role of its DUB domain and related genetic variants remain unclear. We aimed to study the functional effects of A20 DUB-domain alterations in immune cells and understand its link to SLE pathogenesis.Methods: CRISPR/Cas9 was used to generate human U937 monocytes with A20 DUB-inactivating C103A knock-in (KI) mutation. Whole genome RNA-sequencing was used to identify differentially expressed genes between WT and C103A KI cells. Functional studies were performed in A20 C103A U937 cells and in immune cells from A20 C103A mice and genotyped healthy individuals with A20 DUB polymorphism rs2230926. Neutrophil extracellular trap (NET) formation was addressed ex vivo in neutrophils from A20 C103A mice and SLE-patients with rs2230926.Results: Genetic disruption of A20 DUB domain in human and murine myeloid cells did not give rise to enhanced NF-κB signalling. Instead, cells with C103A mutation or rs2230926 polymorphism presented an upregulated expression of PADI4, an enzyme regulating protein citrullination and NET formation, two key mechanisms in autoimmune pathology. A20 C103A cells exhibited enhanced protein citrullination and extracellular trap formation, which could be suppressed by selective PAD4 inhibition. Moreover, SLE-patients with rs2230926 showed increased NETs and increased frequency of autoantibodies to citrullinated epitopes.Conclusions: We propose that genetic alterations disrupting the A20 DUB domain mediate increased susceptibility to SLE through the upregulation of PADI4 with resultant protein citrullination and extracellular trap formation.",

keywords = "NET, PAD4, PADI4, peptidyl arginine deiminase, rs2230926",

author = "Lina Odqvist and Zala Jevnikar and Rebecca Riise and Lisa {\"O}berg and Magdalena Rhedin and Dag Leonard and Linda Yrlid and Sonya Jackson and Johan Mattsson and Sambit Nanda and Philip Cohen and Axel Knebel and Simon Arthur and Kristofer Th{\"o}rn and Elisabet Svenungsson and Andreas J{\"o}nsen and Iva Gunnarsson and Karolina Tandre and Andrei Alexsson and Alf Kastbom and Solbritt Rantap{\"a}{\"a}-Dahlqvist and Maija-Leena Eloranta and Ann-Christine Syv{\"a}nen and Anders Bengtsson and Patrik Johansson and Sandling, {Johanna K.} and Christopher Sj{\"o}wall and Lars R{\"o}nnblom and Barry Collins and Outi Vaarala",

note = "The study was funded by DISSECT AstraZeneca-SciLifeLab collaboration, The Swedish Research Council, Knut and Alice Wallenberg Foundation, Swedish Rheumatism Foundation, King Gustaf V{\textquoteright}s 80-year Foundation, Swedish Society of Medicine, the Ingegerd Johansson donation, and Medical Research Council Programme Grant MRC000985.",

year = "2019",

month = oct,

day = "1",

doi = "10.1136/annrheumdis-2019-215434",

language = "English",

volume = "78",

pages = "1363--1370",

journal = "Annals of the Rheumatic Diseases",

issn = "0003-4967",

publisher = "BMJ Publishing Group",

number = "10",

}

Odqvist, L, Jevnikar, Z, Riise, R, Öberg, L, Rhedin, M, Leonard, D, Yrlid, L, Jackson, S, Mattsson, J, Nanda, S, Cohen, P , Knebel, A , Arthur, S, Thörn, K, Svenungsson, E, Jönsen, A, Gunnarsson, I, Tandre, K, Alexsson, A, Kastbom, A, Rantapää-Dahlqvist, S, Eloranta, M-L, Syvänen, A-C, Bengtsson, A, Johansson, P, Sandling, JK, Sjöwall, C, Rönnblom, L, Collins, B & Vaarala, O 2019, 'Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus', Annals of the Rheumatic Diseases, vol. 78, no. 10, pp. 1363-1370. https://doi.org/10.1136/annrheumdis-2019-215434

TY - JOUR

T1 - Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus

AU - Odqvist, Lina

AU - Jevnikar, Zala

AU - Riise, Rebecca

AU - Öberg, Lisa

AU - Rhedin, Magdalena

AU - Leonard, Dag

AU - Yrlid, Linda

AU - Jackson, Sonya

AU - Mattsson, Johan

AU - Nanda, Sambit

AU - Cohen, Philip

AU - Knebel, Axel

AU - Arthur, Simon

AU - Thörn, Kristofer

AU - Svenungsson, Elisabet

AU - Jönsen, Andreas

AU - Gunnarsson, Iva

AU - Tandre, Karolina

AU - Alexsson, Andrei

AU - Kastbom, Alf

AU - Rantapää-Dahlqvist, Solbritt

AU - Eloranta, Maija-Leena

AU - Syvänen, Ann-Christine

AU - Bengtsson, Anders

AU - Johansson, Patrik

AU - Sandling, Johanna K.

AU - Sjöwall, Christopher

AU - Rönnblom, Lars

AU - Collins, Barry

AU - Vaarala, Outi

N1 - The study was funded by DISSECT AstraZeneca-SciLifeLab collaboration, The Swedish Research Council, Knut and Alice Wallenberg Foundation, Swedish Rheumatism Foundation, King Gustaf V’s 80-year Foundation, Swedish Society of Medicine, the Ingegerd Johansson donation, and Medical Research Council Programme Grant MRC000985.

PY - 2019/10/1

Y1 - 2019/10/1

N2 - Objectives: Genetic variations in TNFAIP3 (A20) de-ubiquitinase (DUB) domain increase the risk of systemic lupus erythematosus (SLE) and rheumatoid arthritis. A20 is a negative regulator of NF-κB but the role of its DUB domain and related genetic variants remain unclear. We aimed to study the functional effects of A20 DUB-domain alterations in immune cells and understand its link to SLE pathogenesis.Methods: CRISPR/Cas9 was used to generate human U937 monocytes with A20 DUB-inactivating C103A knock-in (KI) mutation. Whole genome RNA-sequencing was used to identify differentially expressed genes between WT and C103A KI cells. Functional studies were performed in A20 C103A U937 cells and in immune cells from A20 C103A mice and genotyped healthy individuals with A20 DUB polymorphism rs2230926. Neutrophil extracellular trap (NET) formation was addressed ex vivo in neutrophils from A20 C103A mice and SLE-patients with rs2230926.Results: Genetic disruption of A20 DUB domain in human and murine myeloid cells did not give rise to enhanced NF-κB signalling. Instead, cells with C103A mutation or rs2230926 polymorphism presented an upregulated expression of PADI4, an enzyme regulating protein citrullination and NET formation, two key mechanisms in autoimmune pathology. A20 C103A cells exhibited enhanced protein citrullination and extracellular trap formation, which could be suppressed by selective PAD4 inhibition. Moreover, SLE-patients with rs2230926 showed increased NETs and increased frequency of autoantibodies to citrullinated epitopes.Conclusions: We propose that genetic alterations disrupting the A20 DUB domain mediate increased susceptibility to SLE through the upregulation of PADI4 with resultant protein citrullination and extracellular trap formation.

AB - Objectives: Genetic variations in TNFAIP3 (A20) de-ubiquitinase (DUB) domain increase the risk of systemic lupus erythematosus (SLE) and rheumatoid arthritis. A20 is a negative regulator of NF-κB but the role of its DUB domain and related genetic variants remain unclear. We aimed to study the functional effects of A20 DUB-domain alterations in immune cells and understand its link to SLE pathogenesis.Methods: CRISPR/Cas9 was used to generate human U937 monocytes with A20 DUB-inactivating C103A knock-in (KI) mutation. Whole genome RNA-sequencing was used to identify differentially expressed genes between WT and C103A KI cells. Functional studies were performed in A20 C103A U937 cells and in immune cells from A20 C103A mice and genotyped healthy individuals with A20 DUB polymorphism rs2230926. Neutrophil extracellular trap (NET) formation was addressed ex vivo in neutrophils from A20 C103A mice and SLE-patients with rs2230926.Results: Genetic disruption of A20 DUB domain in human and murine myeloid cells did not give rise to enhanced NF-κB signalling. Instead, cells with C103A mutation or rs2230926 polymorphism presented an upregulated expression of PADI4, an enzyme regulating protein citrullination and NET formation, two key mechanisms in autoimmune pathology. A20 C103A cells exhibited enhanced protein citrullination and extracellular trap formation, which could be suppressed by selective PAD4 inhibition. Moreover, SLE-patients with rs2230926 showed increased NETs and increased frequency of autoantibodies to citrullinated epitopes.Conclusions: We propose that genetic alterations disrupting the A20 DUB domain mediate increased susceptibility to SLE through the upregulation of PADI4 with resultant protein citrullination and extracellular trap formation.

KW - NET

KW - PAD4

KW - PADI4

KW - peptidyl arginine deiminase

KW - rs2230926

UR - http://www.scopus.com/inward/record.url?scp=85069535132&partnerID=8YFLogxK

U2 - 10.1136/annrheumdis-2019-215434

DO - 10.1136/annrheumdis-2019-215434

M3 - Article

C2 - 31300459

SN - 0003-4967

VL - 78

SP - 1363

EP - 1370

JO - Annals of the Rheumatic Diseases

JF - Annals of the Rheumatic Diseases

IS - 10

ER -

Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus

Abstract

Keywords

ASJC Scopus subject areas

UN SDGs

Access to Document

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Elucidation of Mechanisms that Restrict Activation of the Innate Immune System to Prevent Inflammatory and Autoimmune Diseases

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Genetic variations in A20 DUB domain provide a genetic link to citrullination and neutrophil extracellular traps in systemic lupus erythematosus

Abstract

Keywords

ASJC Scopus subject areas

UN SDGs

Access to Document

Other files and links

Fingerprint

Projects

Elucidation of Mechanisms that Restrict Activation of the Innate Immune System to Prevent Inflammatory and Autoimmune Diseases

Cite this