title: C9orf72 promoter hypermethylation is reduced while hydroxymethylation is acquired during reprogramming of ALS patient cells.
abstract: Among several genetic mutations known to cause amyotrophic lateral sclerosis (ALS), a hexanucleotide repeat expansion in the C9orf72 gene is the most common. In approximately 30% of C9orf72-ALS cases, 5-methylcytosine (5mC) levels within the C9orf72 promoter are increased, resulting in a modestly attenuated phenotype. The developmental timing of C9orf72 promoter hypermethylation and the reason why it occurs in only a subset of patients remain unknown. In order to model the acquisition of C9orf72 hypermethylation and examine the potential role of 5-hydroxymethylcytosine (5hmC), we generated induced pluripotent stem cells (iPSCs) from an ALS patient with C9orf72 promoter hypermethylation. Our data show that 5mC levels are reduced by reprogramming and then re-acquired upon neuronal specification, while 5hmC levels increase following reprogramming and are highest in iPSCs and motor neurons. We confirmed the presence of 5hmC within the C9orf72 promoter in post-mortem brain tissues of hypermethylated patients. These findings show that iPSCs are a valuable model system for examining epigenetic perturbations caused by the C9orf72 mutation and reveal a potential role for cytosine demethylation.
authors.affiliation: Center for Therapeutic Innovation, University of Miami Miller School of Medicine, Miami, FL, USA; Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL, USA.
authors.forename: Rustam
authors.initials: R
authors.lastname: Esanov
authors.affiliation: John P. Hussman Institute for Human Genomics, University of Miami Miller School of Medicine, USA; The Dr. John T. Macdonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, USA.
authors.forename: Kinsley C
authors.initials: KC
authors.lastname: Belle
authors.affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA.
authors.forename: Marka
authors.initials: M
authors.lastname: van Blitterswijk
authors.affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA.
authors.forename: Veronique V
authors.initials: VV
authors.lastname: Belzil
authors.affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA.
authors.forename: Rosa
authors.initials: R
authors.lastname: Rademakers
authors.affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA.
authors.forename: Dennis W
authors.initials: DW
authors.lastname: Dickson
authors.affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA.
authors.forename: Leonard
authors.initials: L
authors.lastname: Petrucelli
authors.affiliation: Department of Neurology, Mayo Clinic, Jacksonville, FL, USA.
authors.forename: Kevin B
authors.initials: KB
authors.lastname: Boylan
authors.affiliation: John P. Hussman Institute for Human Genomics, University of Miami Miller School of Medicine, USA; The Dr. John T. Macdonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, USA.
authors.forename: Derek M
authors.initials: DM
authors.lastname: Dykxhoorn
authors.affiliation: Department of Neurology, University of Miami Miller School of Medicine, USA.
authors.forename: Joanne
authors.initials: J
authors.lastname: Wuu
authors.affiliation: Department of Neurology, University of Miami Miller School of Medicine, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Benatar
authors.affiliation: Center for Therapeutic Innovation, University of Miami Miller School of Medicine, Miami, FL, USA; Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL, USA.
authors.forename: Claes
authors.initials: C
authors.lastname: Wahlestedt
authors.affiliation: Center for Therapeutic Innovation, University of Miami Miller School of Medicine, Miami, FL, USA; Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL, USA. Electronic address: zzeier@med.miami.edu.
authors.forename: Zane
authors.initials: Z
authors.lastname: Zeier
mesh_terms: D044503:5-Methylcytosine / Q000378:metabolism / Q000031:analogs & derivatives; D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics* / Q000473:pathology*; D001921:Brain / Q000473:pathology; D000073885:C9orf72 Protein; D018920:Coculture Techniques; D018899:CpG Islands / Q000502:physiology; D003596:Cytosine / Q000031:analogs & derivatives; D019175:DNA Methylation / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000502:physiology*; D008214:Lymphocytes / Q000502:physiology; D009046:Motor Neurons / Q000502:physiology; D009154:Mutation / Q000235:genetics*; D000071317:Nanog Homeobox Protein; D064231:Nestin / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000378:metabolism; D013997:Time Factors
chemical_list: D000073885:C9orf72 Protein; C568651:C9orf72 protein, human; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D064231:Nestin; D011506:Proteins; D012333:RNA, Messenger; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; C011865:5-hydroxymethylcytosine; D049452:Green Fluorescent Proteins; D044503:5-Methylcytosine; D003596:Cytosine
keywords: Amyotrophic lateral sclerosis; C9orf72; Cytosine hydroxymethylation; Repeat expansion