title: TDP-43 and FUS RNA-binding proteins bind distinct sets of cytoplasmic messenger RNAs and differently regulate their post-transcriptional fate in motoneuron-like cells.
abstract: The RNA-binding proteins TDP-43 and FUS form abnormal cytoplasmic aggregates in affected tissues of patients with amyotrophic lateral sclerosis and frontotemporal lobar dementia. TDP-43 and FUS localize mainly in the nucleus where they regulate pre-mRNA splicing, but they are also involved in mRNA transport, stability, and translation. To better investigate their cytoplasmic activities, we applied an RNA immunoprecipitation and chip analysis to define the mRNAs associated to TDP-43 and FUS in the cytoplasmic ribonucleoprotein complexes from motoneuronal NSC-34 cells. We found that they bind different sets of mRNAs although converging on common cellular pathways. Bioinformatics analyses identified the (UG)(n) consensus motif in 80% of 3'-UTR sequences of TDP-43 targets, whereas for FUS the binding motif was less evident. By in vitro assays we validated binding to selected target 3'-UTRs, including Vegfa and Grn for TDP-43, and Vps54, Nvl, and Taf15 for FUS. We showed that TDP-43 has a destabilizing activity on Vegfa and Grn mRNAs and may ultimately affect progranulin protein content, whereas FUS does not affect mRNA stability/translation of its targets. We also demonstrated that three different point mutations in TDP-43 did not change the binding affinity for Vegfa and Grn mRNAs or their protein level. Our data indicate that TDP-43 and FUS recognize distinct sets of mRNAs and differently regulate their fate in the cytoplasm of motoneuron-like cells, therefore suggesting complementary roles in neuronal RNA metabolism and neurodegeneration.
authors.affiliation: Department of Neurology and Laboratory of Neuroscience, IRCCS Istituto Auxologico Italiano, Milan 20149, Italy.
authors.forename: Claudia
authors.initials: C
authors.lastname: Colombrita
authors.forename: Elisa
authors.initials: E
authors.lastname: Onesto
authors.forename: Francesca
authors.initials: F
authors.lastname: Megiorni
authors.forename: Antonio
authors.initials: A
authors.lastname: Pizzuti
authors.forename: Francisco E
authors.initials: FE
authors.lastname: Baralle
authors.forename: Emanuele
authors.initials: E
authors.lastname: Buratti
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Silani
authors.forename: Antonia
authors.initials: A
authors.lastname: Ratti
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000077155:Granulins; D006801:Humans; D047468:Immunoprecipitation; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009046:Motor Neurons / Q000166:cytology / Q000378:metabolism*; D009154:Mutation; D059372:Nucleotide Motifs / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D000077153:Progranulins; D011485:Protein Binding; D014176:Protein Biosynthesis; D034622:RNA Interference; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D034702:RNA-Binding Protein FUS / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D035381:TATA-Binding Protein Associated Factors / Q000235:genetics / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D020413:3' Untranslated Regions; D004268:DNA-Binding Proteins; D000077155:Granulins; C120204:Grn protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D000077153:Progranulins; D012333:RNA, Messenger; D034702:RNA-Binding Protein FUS; D035381:TATA-Binding Protein Associated Factors; D042461:Vascular Endothelial Growth Factor A; D033921:Vesicular Transport Proteins; C486750:Vps54 protein, mouse