title: MTHFSD and DDX58 are novel RNA-binding proteins abnormally regulated in amyotrophic lateral sclerosis.
abstract: Tar DNA-binding protein 43 (TDP-43) is an RNA-binding protein normally localized to the nucleus of cells, where it elicits functions related to RNA metabolism such as transcriptional regulation and alternative splicing. In amyotrophic lateral sclerosis, TDP-43 is mislocalized from the nucleus to the cytoplasm of diseased motor neurons, forming ubiquitinated inclusions. Although mutations in the gene encoding TDP-43, TARDBP, are found in amyotrophic lateral sclerosis, these are rare. However, TDP-43 pathology is common to over 95% of amyotrophic lateral sclerosis cases, suggesting that abnormalities of TDP-43 play an active role in disease pathogenesis. It is our hypothesis that a loss of TDP-43 from the nucleus of affected motor neurons in amyotrophic lateral sclerosis will lead to changes in RNA processing and expression. Identifying these changes could uncover molecular pathways that underpin motor neuron degeneration. Here we have used translating ribosome affinity purification coupled with microarray analysis to identify the mRNAs being actively translated in motor neurons of mutant TDP-43(A315T) mice compared to age-matched non-transgenic littermates. No significant changes were found at 5 months (presymptomatic) of age, but at 10 months (symptomatic) the translational profile revealed significant changes in genes involved in RNA metabolic process, immune response and cell cycle regulation. Of 28 differentially expressed genes, seven had a ≥ 2-fold change; four were validated by immunofluorescence labelling of motor neurons in TDP-43(A315T) mice, and two of these were confirmed by immunohistochemistry in amyotrophic lateral sclerosis cases. Both of these identified genes, DDX58 and MTHFSD, are RNA-binding proteins, and we show that TDP-43 binds to their respective mRNAs and we identify MTHFSD as a novel component of stress granules. This discovery-based approach has for the first time revealed translational changes in motor neurons of a TDP-43 mouse model, identifying DDX58 and MTHFSD as two TDP-43 targets that are misregulated in amyotrophic lateral sclerosis.
authors.affiliation: 1 Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, M5T 2S8, Canada 2 Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, M5S 1A1, Canada.
authors.forename: Laura
authors.initials: L
authors.lastname: MacNair
authors.affiliation: 1 Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, M5T 2S8, Canada.
authors.forename: Shangxi
authors.initials: S
authors.lastname: Xiao
authors.affiliation: 1 Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, M5T 2S8, Canada.
authors.forename: Denise
authors.initials: D
authors.lastname: Miletic
authors.affiliation: 1 Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, M5T 2S8, Canada.
authors.forename: Mahdi
authors.initials: M
authors.lastname: Ghani
authors.affiliation: 3 Département de psychiatrie et de neurosciences, Université Laval, Québec G1V 0A6, Canada.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Julien
authors.affiliation: 2 Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, M5S 1A1, Canada 4 Sunnybrook Health Sciences Centre, 2075 Bayview Avenue, Toronto, ON, M4N 3M5, Canada.
authors.forename: Julia
authors.initials: J
authors.lastname: Keith
authors.affiliation: 4 Sunnybrook Health Sciences Centre, 2075 Bayview Avenue, Toronto, ON, M4N 3M5, Canada.
authors.forename: Lorne
authors.initials: L
authors.lastname: Zinman
authors.affiliation: 1 Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, M5T 2S8, Canada.
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Rogaeva
authors.affiliation: 1 Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, M5T 2S8, Canada 2 Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, M5S 1A1, Canada jan.robertson@utoronto.ca.
authors.forename: Janice
authors.initials: J
authors.lastname: Robertson
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics; D000818:Animals; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics*; D005786:Gene Expression Regulation / Q000235:genetics*; D006801:Humans; D051379:Mice; D009154:Mutation; D016601:RNA-Binding Proteins / Q000235:genetics*
chemical_list: D004268:DNA-Binding Proteins; D016601:RNA-Binding Proteins; C503356:Ddx58 protein, mouse; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases
keywords: ALS; RNA-binding proteins; TDP-43; translational profiling