title: RAN proteins and RNA foci from antisense transcripts in C9ORF72 ALS and frontotemporal dementia.
abstract: The finding that a GGGGCC (G4C2) hexanucleotide repeat expansion in the chromosome 9 ORF 72 (C9ORF72) gene is a common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) links ALS/FTD to a large group of unstable microsatellite diseases. Previously, we showed that microsatellite expansion mutations can be bidirectionally transcribed and that these mutations express unexpected proteins by a unique mechanism, repeat-associated non-ATG (RAN) translation. In this study, we show that C9ORF72 antisense transcripts are elevated in the brains of C9ORF72 expansion-positive [C9(+)] patients, and antisense GGCCCC (G2C4) repeat-expansion RNAs accumulate in nuclear foci in brain. Additionally, sense and antisense foci accumulate in blood and are potential biomarkers of the disease. Furthermore, we show that RAN translation occurs from both sense and antisense expansion transcripts, resulting in the expression of six RAN proteins (antisense: Pro-Arg, Pro-Ala, Gly-Pro; and sense: Gly-Ala, Gly-Arg, Gly-Pro). These proteins accumulate in cytoplasmic aggregates in affected brain regions, including the frontal and motor cortex, hippocampus, and spinal cord neurons, with some brain regions showing dramatic RAN protein accumulation and clustering. The finding that unique antisense G2C4 RNA foci and three unique antisense RAN proteins accumulate in patient tissues indicates that bidirectional transcription of expanded alleles is a fundamental pathologic feature of C9ORF72 ALS/FTD. Additionally, these findings suggest the need to test therapeutic strategies that target both sense and antisense RNAs and RAN proteins in C9ORF72 ALS/FTD, and to more broadly consider the role of antisense expression and RAN translation across microsatellite expansion diseases.
authors.affiliation: Department of Molecular Genetics and Microbiology, Department of Neurology, and Department of Neuroscience, Center for NeuroGenetics, Genetics Institute, College of Medicine, University of Florida, Gainesville, FL 32610.
authors.forename: Tao
authors.initials: T
authors.lastname: Zu
authors.forename: Yuanjing
authors.initials: Y
authors.lastname: Liu
authors.forename: Monica
authors.initials: M
authors.lastname: Bañez-Coronel
authors.forename: Tammy
authors.initials: T
authors.lastname: Reid
authors.forename: Olga
authors.initials: O
authors.lastname: Pletnikova
authors.forename: Jada
authors.initials: J
authors.lastname: Lewis
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Miller
authors.forename: Matthew B
authors.initials: MB
authors.lastname: Harms
authors.forename: Annet E
authors.initials: AE
authors.lastname: Falchook
authors.forename: S H
authors.initials: SH
authors.lastname: Subramony
authors.forename: Lyle W
authors.initials: LW
authors.lastname: Ostrow
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Rothstein
authors.forename: Juan C
authors.initials: JC
authors.lastname: Troncoso
authors.forename: Laura P W
authors.initials: LP
authors.lastname: Ranum
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D001921:Brain / Q000378:metabolism* / Q000473:pathology; D000073885:C9orf72 Protein; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005260:Female; D057180:Frontotemporal Dementia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D016372:RNA, Antisense / Q000096:biosynthesis* / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D020931:ran GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D000073885:C9orf72 Protein; C568651:C9orf72 protein, human; D009419:Nerve Tissue Proteins; D011506:Proteins; C120587:RAN protein, human; D016372:RNA, Antisense; D020931:ran GTP-Binding Protein
keywords: clustered aggregates; cytoplasmic inclusions; noncoding RNA