Identification of new genetic syndromes with epilepsy by whole-exome sequencing

Mikko Muona

Research output: ThesisDoctoral ThesisCollection of Articles

Abstract

Epilepsies are a heterogeneous group of central nervous system diseases characterised by recurrent epileptic seizures. They are one of the most common neurological diseases with a lifetime prevalence of ~4%. Epileptic seizures are also a common comorbidity of various neurobiological disorders where epilepsy is not the primary diagnosis. Most epilepsies have a genetic origin, either monogenic or polygenic, however, the causal genetic variants have remained unknown in a substantial proportion of individuals with epilepsies. Over the past decade, technological advances in DNA sequencing have allowed the characterisation of the genetic basis of human disorders rapidly and efficiently. One of the most widely used methods is whole-exome sequencing (WES) where genetic variants in the protein coding regions of the genome, the exome, are captured. Even though the exome constitutes only ~1.5% of the genome, the majority of disease-causing variants underlying severe, monogenic diseases are located in the protein coding regions. Here, we aimed to decipher the molecular genetic basis of severe epilepsy syndromes by utilising WES to identify disease-causing genetic variants in patients without a genetic diagnosis. We studied patients with progressive myoclonus epilepsy (PME, n=84) or severe infantile-onset epileptic syndromes (n=30), which are one of the most devastating forms of genetic syndromes with epilepsy and characterised by frequent, pharmacoresistant seizures and poor prognosis. Given that the patients had undergone genetic testing to varying extent prior to this study, we specifically aimed to establish novel genes and molecular biological mechanisms underlying these syndromes. We made substantial progress in understanding the genetic architecture and molecular basis of the studied syndromes. For PMEs, we established a new major genetic cause and also expanded the genotypic and phenotypic spectrum of previously established disease genes. For severe infantile-onset epileptic syndromes, we identified one new, definite causal gene and one that requires identification of additional patients to confirm the causal role. The three newly identified disease genes represent three different molecular functions that together give new insight on epileptogenic mechanisms. The new PME subtype is caused by a heterozygous missense variant c.959G>A (p.Arg320His) in KCNC1 that was identified in 11 unrelated patients (13%) in the PME exome sequencing cohort. We have subsequently identified six additional patients. The gene encodes a potassium ion channel KV3.1 that has an important role in generating action potentials in the central nervous system, with the mutation disrupting the ability to transport potassium ions across the cell membrane. This mutation occurs in most families de novo, that is, it is a newly arising mutation. Based on the estimated mutation rate, the recurrent KCNC1 mutation is a worldwide cause of PME with likely hundreds of affected individuals globally. In five families with altogether nine affected siblings, we identified compound heterozygous variants in UBA5 as the cause of an infantile-onset syndrome characterised initially by irritability, followed by epilepsy, dystonic movements, moderate to severe intellectual disability, microcephaly and stagnation of development. The gene encodes an activating enzyme for UFM1, which is a small ubiquitin-like protein that is conjugated to its target proteins. The function of the highly conserved UFM1 conjugation system is still largely unknown. Functional analysis of the UBA5 mutants suggest that the identified variants cause reduced enzymatic activity of UBA5. Symptoms of the UBA5 patients and our findings in the central nervous system specific knockout mice for Ufm1 together indicate that UFM1-cascade is essential for normal development and function of the central nervous system. Finally, we identified compound heterozygous variants in ADAM22 as the likely cause of the disease in a patient with an infantile-onset rapidly progressing encephalopathy with epilepsy and cortical atrophy. The gene encodes a postsynaptic protein that functions as a receptor for LGI1, and we show that the identified variants abolish the ability of ADAM22 to bind to LGI1. The LGI1-ADAM22 complex is an antiepileptogenic factor regulating synaptic transmission throughout life. Highlighting the important role of this complex, knockout of Adam22 and Lgi1 in mice causes lethal epilepsy. Autosomal dominant LGI1 variants also cause epilepsy in humans. Identification of a patient with loss-of-function variants in ADAM22 suggest that also this gene is linked to epilepsy in humans. This connection should be confirmed through identification of additional affected individuals with ADAM22 variants. Altogether, this thesis demonstrates the power of WES in identification of causal genetic variants even in phenotypically heterogeneous patient cohorts subjected to prior genetic screenings. The findings improve diagnostics of these syndromes, increase knowledge of the underlying molecular mechanisms and potentially aid in developing new therapeutic interventions. Finally, for these families, establishment of the genetic diagnosis ends years of uncertainty and frustration of not knowing the cause of the disease and prevents need for unnecessary diagnostic testing.Epilepsiat ovat heterogeeninen joukko keskushermostosairauksia, jotka ilmenevät toistuvina epileptisinä kohtauksina. Niiden elämänaikainen esiintyvyys on noin 4 % eli ne ovat yksiä yleisimmistä neurologisista sairauksista. Epileptisiä kohtauksia esiintyy myös osana muita keskushermoston sairauksia, joissa epilepsia ei ole päädiagnoosi. Useimmat epilepsiat ovat geneettisiä joko mono- tai polygeenisiä mutta tautia aiheuttavat geneettiset variantit jäävät tunnistamatta merkittävällä osalla epilepsiaa sairastavista.
Original languageEnglish
Place of PublicationHelsinki
Publisher
Print ISBNs978-951-51-2255-1
Electronic ISBNs978-951-51-2256-8
Publication statusPublished - 2016
MoE publication typeG5 Doctoral dissertation (article)

Fields of Science

  • 3112 Neurosciences
  • 3111 Biomedicine

Cite this

Muona, M. (2016). Identification of new genetic syndromes with epilepsy by whole-exome sequencing. Helsinki: University of Helsinki.
Muona, Mikko. / Identification of new genetic syndromes with epilepsy by whole-exome sequencing. Helsinki : University of Helsinki, 2016. 117 p.
@phdthesis{5eb41622b6f14e848200e8ab6a51e4ee,
title = "Identification of new genetic syndromes with epilepsy by whole-exome sequencing",
abstract = "Epilepsies are a heterogeneous group of central nervous system diseases characterised by recurrent epileptic seizures. They are one of the most common neurological diseases with a lifetime prevalence of ~4{\%}. Epileptic seizures are also a common comorbidity of various neurobiological disorders where epilepsy is not the primary diagnosis. Most epilepsies have a genetic origin, either monogenic or polygenic, however, the causal genetic variants have remained unknown in a substantial proportion of individuals with epilepsies. Over the past decade, technological advances in DNA sequencing have allowed the characterisation of the genetic basis of human disorders rapidly and efficiently. One of the most widely used methods is whole-exome sequencing (WES) where genetic variants in the protein coding regions of the genome, the exome, are captured. Even though the exome constitutes only ~1.5{\%} of the genome, the majority of disease-causing variants underlying severe, monogenic diseases are located in the protein coding regions. Here, we aimed to decipher the molecular genetic basis of severe epilepsy syndromes by utilising WES to identify disease-causing genetic variants in patients without a genetic diagnosis. We studied patients with progressive myoclonus epilepsy (PME, n=84) or severe infantile-onset epileptic syndromes (n=30), which are one of the most devastating forms of genetic syndromes with epilepsy and characterised by frequent, pharmacoresistant seizures and poor prognosis. Given that the patients had undergone genetic testing to varying extent prior to this study, we specifically aimed to establish novel genes and molecular biological mechanisms underlying these syndromes. We made substantial progress in understanding the genetic architecture and molecular basis of the studied syndromes. For PMEs, we established a new major genetic cause and also expanded the genotypic and phenotypic spectrum of previously established disease genes. For severe infantile-onset epileptic syndromes, we identified one new, definite causal gene and one that requires identification of additional patients to confirm the causal role. The three newly identified disease genes represent three different molecular functions that together give new insight on epileptogenic mechanisms. The new PME subtype is caused by a heterozygous missense variant c.959G>A (p.Arg320His) in KCNC1 that was identified in 11 unrelated patients (13{\%}) in the PME exome sequencing cohort. We have subsequently identified six additional patients. The gene encodes a potassium ion channel KV3.1 that has an important role in generating action potentials in the central nervous system, with the mutation disrupting the ability to transport potassium ions across the cell membrane. This mutation occurs in most families de novo, that is, it is a newly arising mutation. Based on the estimated mutation rate, the recurrent KCNC1 mutation is a worldwide cause of PME with likely hundreds of affected individuals globally. In five families with altogether nine affected siblings, we identified compound heterozygous variants in UBA5 as the cause of an infantile-onset syndrome characterised initially by irritability, followed by epilepsy, dystonic movements, moderate to severe intellectual disability, microcephaly and stagnation of development. The gene encodes an activating enzyme for UFM1, which is a small ubiquitin-like protein that is conjugated to its target proteins. The function of the highly conserved UFM1 conjugation system is still largely unknown. Functional analysis of the UBA5 mutants suggest that the identified variants cause reduced enzymatic activity of UBA5. Symptoms of the UBA5 patients and our findings in the central nervous system specific knockout mice for Ufm1 together indicate that UFM1-cascade is essential for normal development and function of the central nervous system. Finally, we identified compound heterozygous variants in ADAM22 as the likely cause of the disease in a patient with an infantile-onset rapidly progressing encephalopathy with epilepsy and cortical atrophy. The gene encodes a postsynaptic protein that functions as a receptor for LGI1, and we show that the identified variants abolish the ability of ADAM22 to bind to LGI1. The LGI1-ADAM22 complex is an antiepileptogenic factor regulating synaptic transmission throughout life. Highlighting the important role of this complex, knockout of Adam22 and Lgi1 in mice causes lethal epilepsy. Autosomal dominant LGI1 variants also cause epilepsy in humans. Identification of a patient with loss-of-function variants in ADAM22 suggest that also this gene is linked to epilepsy in humans. This connection should be confirmed through identification of additional affected individuals with ADAM22 variants. Altogether, this thesis demonstrates the power of WES in identification of causal genetic variants even in phenotypically heterogeneous patient cohorts subjected to prior genetic screenings. The findings improve diagnostics of these syndromes, increase knowledge of the underlying molecular mechanisms and potentially aid in developing new therapeutic interventions. Finally, for these families, establishment of the genetic diagnosis ends years of uncertainty and frustration of not knowing the cause of the disease and prevents need for unnecessary diagnostic testing.Epilepsiat ovat heterogeeninen joukko keskushermostosairauksia, jotka ilmenev{\"a}t toistuvina epileptisin{\"a} kohtauksina. Niiden el{\"a}m{\"a}naikainen esiintyvyys on noin 4 {\%} eli ne ovat yksi{\"a} yleisimmist{\"a} neurologisista sairauksista. Epileptisi{\"a} kohtauksia esiintyy my{\"o}s osana muita keskushermoston sairauksia, joissa epilepsia ei ole p{\"a}{\"a}diagnoosi. Useimmat epilepsiat ovat geneettisi{\"a} joko mono- tai polygeenisi{\"a} mutta tautia aiheuttavat geneettiset variantit j{\"a}{\"a}v{\"a}t tunnistamatta merkitt{\"a}v{\"a}ll{\"a} osalla epilepsiaa sairastavista.",
keywords = "ADAM Proteins, +genetics, Brain Diseases, DNA Mutational Analysis, Epilepsy, Exome, Molecular Sequence Data, Mutation, Missense, Myoclonic Epilepsies, Progressive, Nerve Tissue Proteins, Point Mutation, Sequence Analysis, DNA, Shaw Potassium Channels, Ubiquitin-Activating Enzymes, 3112 Neurosciences, 3111 Biomedicine",
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Identification of new genetic syndromes with epilepsy by whole-exome sequencing. / Muona, Mikko.

Helsinki : University of Helsinki, 2016. 117 p.

Research output: ThesisDoctoral ThesisCollection of Articles

TY - THES

T1 - Identification of new genetic syndromes with epilepsy by whole-exome sequencing

AU - Muona, Mikko

N1 - M1 - 117 s. + liitteet Helsingin yliopisto Volume: Proceeding volume:

PY - 2016

Y1 - 2016

N2 - Epilepsies are a heterogeneous group of central nervous system diseases characterised by recurrent epileptic seizures. They are one of the most common neurological diseases with a lifetime prevalence of ~4%. Epileptic seizures are also a common comorbidity of various neurobiological disorders where epilepsy is not the primary diagnosis. Most epilepsies have a genetic origin, either monogenic or polygenic, however, the causal genetic variants have remained unknown in a substantial proportion of individuals with epilepsies. Over the past decade, technological advances in DNA sequencing have allowed the characterisation of the genetic basis of human disorders rapidly and efficiently. One of the most widely used methods is whole-exome sequencing (WES) where genetic variants in the protein coding regions of the genome, the exome, are captured. Even though the exome constitutes only ~1.5% of the genome, the majority of disease-causing variants underlying severe, monogenic diseases are located in the protein coding regions. Here, we aimed to decipher the molecular genetic basis of severe epilepsy syndromes by utilising WES to identify disease-causing genetic variants in patients without a genetic diagnosis. We studied patients with progressive myoclonus epilepsy (PME, n=84) or severe infantile-onset epileptic syndromes (n=30), which are one of the most devastating forms of genetic syndromes with epilepsy and characterised by frequent, pharmacoresistant seizures and poor prognosis. Given that the patients had undergone genetic testing to varying extent prior to this study, we specifically aimed to establish novel genes and molecular biological mechanisms underlying these syndromes. We made substantial progress in understanding the genetic architecture and molecular basis of the studied syndromes. For PMEs, we established a new major genetic cause and also expanded the genotypic and phenotypic spectrum of previously established disease genes. For severe infantile-onset epileptic syndromes, we identified one new, definite causal gene and one that requires identification of additional patients to confirm the causal role. The three newly identified disease genes represent three different molecular functions that together give new insight on epileptogenic mechanisms. The new PME subtype is caused by a heterozygous missense variant c.959G>A (p.Arg320His) in KCNC1 that was identified in 11 unrelated patients (13%) in the PME exome sequencing cohort. We have subsequently identified six additional patients. The gene encodes a potassium ion channel KV3.1 that has an important role in generating action potentials in the central nervous system, with the mutation disrupting the ability to transport potassium ions across the cell membrane. This mutation occurs in most families de novo, that is, it is a newly arising mutation. Based on the estimated mutation rate, the recurrent KCNC1 mutation is a worldwide cause of PME with likely hundreds of affected individuals globally. In five families with altogether nine affected siblings, we identified compound heterozygous variants in UBA5 as the cause of an infantile-onset syndrome characterised initially by irritability, followed by epilepsy, dystonic movements, moderate to severe intellectual disability, microcephaly and stagnation of development. The gene encodes an activating enzyme for UFM1, which is a small ubiquitin-like protein that is conjugated to its target proteins. The function of the highly conserved UFM1 conjugation system is still largely unknown. Functional analysis of the UBA5 mutants suggest that the identified variants cause reduced enzymatic activity of UBA5. Symptoms of the UBA5 patients and our findings in the central nervous system specific knockout mice for Ufm1 together indicate that UFM1-cascade is essential for normal development and function of the central nervous system. Finally, we identified compound heterozygous variants in ADAM22 as the likely cause of the disease in a patient with an infantile-onset rapidly progressing encephalopathy with epilepsy and cortical atrophy. The gene encodes a postsynaptic protein that functions as a receptor for LGI1, and we show that the identified variants abolish the ability of ADAM22 to bind to LGI1. The LGI1-ADAM22 complex is an antiepileptogenic factor regulating synaptic transmission throughout life. Highlighting the important role of this complex, knockout of Adam22 and Lgi1 in mice causes lethal epilepsy. Autosomal dominant LGI1 variants also cause epilepsy in humans. Identification of a patient with loss-of-function variants in ADAM22 suggest that also this gene is linked to epilepsy in humans. This connection should be confirmed through identification of additional affected individuals with ADAM22 variants. Altogether, this thesis demonstrates the power of WES in identification of causal genetic variants even in phenotypically heterogeneous patient cohorts subjected to prior genetic screenings. The findings improve diagnostics of these syndromes, increase knowledge of the underlying molecular mechanisms and potentially aid in developing new therapeutic interventions. Finally, for these families, establishment of the genetic diagnosis ends years of uncertainty and frustration of not knowing the cause of the disease and prevents need for unnecessary diagnostic testing.Epilepsiat ovat heterogeeninen joukko keskushermostosairauksia, jotka ilmenevät toistuvina epileptisinä kohtauksina. Niiden elämänaikainen esiintyvyys on noin 4 % eli ne ovat yksiä yleisimmistä neurologisista sairauksista. Epileptisiä kohtauksia esiintyy myös osana muita keskushermoston sairauksia, joissa epilepsia ei ole päädiagnoosi. Useimmat epilepsiat ovat geneettisiä joko mono- tai polygeenisiä mutta tautia aiheuttavat geneettiset variantit jäävät tunnistamatta merkittävällä osalla epilepsiaa sairastavista.

AB - Epilepsies are a heterogeneous group of central nervous system diseases characterised by recurrent epileptic seizures. They are one of the most common neurological diseases with a lifetime prevalence of ~4%. Epileptic seizures are also a common comorbidity of various neurobiological disorders where epilepsy is not the primary diagnosis. Most epilepsies have a genetic origin, either monogenic or polygenic, however, the causal genetic variants have remained unknown in a substantial proportion of individuals with epilepsies. Over the past decade, technological advances in DNA sequencing have allowed the characterisation of the genetic basis of human disorders rapidly and efficiently. One of the most widely used methods is whole-exome sequencing (WES) where genetic variants in the protein coding regions of the genome, the exome, are captured. Even though the exome constitutes only ~1.5% of the genome, the majority of disease-causing variants underlying severe, monogenic diseases are located in the protein coding regions. Here, we aimed to decipher the molecular genetic basis of severe epilepsy syndromes by utilising WES to identify disease-causing genetic variants in patients without a genetic diagnosis. We studied patients with progressive myoclonus epilepsy (PME, n=84) or severe infantile-onset epileptic syndromes (n=30), which are one of the most devastating forms of genetic syndromes with epilepsy and characterised by frequent, pharmacoresistant seizures and poor prognosis. Given that the patients had undergone genetic testing to varying extent prior to this study, we specifically aimed to establish novel genes and molecular biological mechanisms underlying these syndromes. We made substantial progress in understanding the genetic architecture and molecular basis of the studied syndromes. For PMEs, we established a new major genetic cause and also expanded the genotypic and phenotypic spectrum of previously established disease genes. For severe infantile-onset epileptic syndromes, we identified one new, definite causal gene and one that requires identification of additional patients to confirm the causal role. The three newly identified disease genes represent three different molecular functions that together give new insight on epileptogenic mechanisms. The new PME subtype is caused by a heterozygous missense variant c.959G>A (p.Arg320His) in KCNC1 that was identified in 11 unrelated patients (13%) in the PME exome sequencing cohort. We have subsequently identified six additional patients. The gene encodes a potassium ion channel KV3.1 that has an important role in generating action potentials in the central nervous system, with the mutation disrupting the ability to transport potassium ions across the cell membrane. This mutation occurs in most families de novo, that is, it is a newly arising mutation. Based on the estimated mutation rate, the recurrent KCNC1 mutation is a worldwide cause of PME with likely hundreds of affected individuals globally. In five families with altogether nine affected siblings, we identified compound heterozygous variants in UBA5 as the cause of an infantile-onset syndrome characterised initially by irritability, followed by epilepsy, dystonic movements, moderate to severe intellectual disability, microcephaly and stagnation of development. The gene encodes an activating enzyme for UFM1, which is a small ubiquitin-like protein that is conjugated to its target proteins. The function of the highly conserved UFM1 conjugation system is still largely unknown. Functional analysis of the UBA5 mutants suggest that the identified variants cause reduced enzymatic activity of UBA5. Symptoms of the UBA5 patients and our findings in the central nervous system specific knockout mice for Ufm1 together indicate that UFM1-cascade is essential for normal development and function of the central nervous system. Finally, we identified compound heterozygous variants in ADAM22 as the likely cause of the disease in a patient with an infantile-onset rapidly progressing encephalopathy with epilepsy and cortical atrophy. The gene encodes a postsynaptic protein that functions as a receptor for LGI1, and we show that the identified variants abolish the ability of ADAM22 to bind to LGI1. The LGI1-ADAM22 complex is an antiepileptogenic factor regulating synaptic transmission throughout life. Highlighting the important role of this complex, knockout of Adam22 and Lgi1 in mice causes lethal epilepsy. Autosomal dominant LGI1 variants also cause epilepsy in humans. Identification of a patient with loss-of-function variants in ADAM22 suggest that also this gene is linked to epilepsy in humans. This connection should be confirmed through identification of additional affected individuals with ADAM22 variants. Altogether, this thesis demonstrates the power of WES in identification of causal genetic variants even in phenotypically heterogeneous patient cohorts subjected to prior genetic screenings. The findings improve diagnostics of these syndromes, increase knowledge of the underlying molecular mechanisms and potentially aid in developing new therapeutic interventions. Finally, for these families, establishment of the genetic diagnosis ends years of uncertainty and frustration of not knowing the cause of the disease and prevents need for unnecessary diagnostic testing.Epilepsiat ovat heterogeeninen joukko keskushermostosairauksia, jotka ilmenevät toistuvina epileptisinä kohtauksina. Niiden elämänaikainen esiintyvyys on noin 4 % eli ne ovat yksiä yleisimmistä neurologisista sairauksista. Epileptisiä kohtauksia esiintyy myös osana muita keskushermoston sairauksia, joissa epilepsia ei ole päädiagnoosi. Useimmat epilepsiat ovat geneettisiä joko mono- tai polygeenisiä mutta tautia aiheuttavat geneettiset variantit jäävät tunnistamatta merkittävällä osalla epilepsiaa sairastavista.

KW - ADAM Proteins

KW - +genetics

KW - Brain Diseases

KW - DNA Mutational Analysis

KW - Epilepsy

KW - Exome

KW - Molecular Sequence Data

KW - Mutation, Missense

KW - Myoclonic Epilepsies, Progressive

KW - Nerve Tissue Proteins

KW - Point Mutation

KW - Sequence Analysis, DNA

KW - Shaw Potassium Channels

KW - Ubiquitin-Activating Enzymes

KW - 3112 Neurosciences

KW - 3111 Biomedicine

M3 - Doctoral Thesis

SN - 978-951-51-2255-1

T3 - Dissertationes Scholae Doctoralis Ad Sanitatem Investigandam Universitatis Helsinkiensis

PB - University of Helsinki

CY - Helsinki

ER -

Muona M. Identification of new genetic syndromes with epilepsy by whole-exome sequencing. Helsinki: University of Helsinki, 2016. 117 p. (Dissertationes Scholae Doctoralis Ad Sanitatem Investigandam Universitatis Helsinkiensis; 44/2016).