The Identification of Nuclear FMRP Isoform Iso6 Partners
Author:
Ledoux Nassim1, Lelong Emeline I. J.1ORCID, Simard Alexandre1, Hussein Samer1, Adjibade Pauline1, Lambert Jean-Philippe23ORCID, Mazroui Rachid1
Affiliation:
1. Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada 2. Centre de Recherche du CHU de Québec—Université Laval, Axe Endocrinologie et néphrologie, Département de Médecine Moléculaire, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada 3. PROTEO, Le Regroupement Québécois De Recherche Sur La Fonction, L’ingénierie et Les Applications des Protéines, Université Laval, Québec, QC G1V 0A6, Canada
Abstract
A deficiency of FMRP, a canonical RNA-binding protein, causes the development of Fragile X Syndrome (FXS), which is characterised by multiple phenotypes, including neurodevelopmental disorders, intellectual disability, and autism. Due to the alternative splicing of the encoding FMR1 gene, multiple FMRP isoforms are produced consisting of full-length predominantly cytoplasmic (i.e., iso1) isoforms involved in translation and truncated nuclear (i.e., iso6) isoforms with orphan functions. However, we recently implicated nuclear FMRP isoforms in DNA damage response, showing that they negatively regulate the accumulation of anaphase DNA genomic instability bridges. This finding provided evidence that the cytoplasmic and nuclear functions of FMRP are uncoupled played by respective cytoplasmic and nuclear isoforms, potentially involving specific interactions. While interaction partners of cytoplasmic FMRP have been reported, the identity of nuclear FMRP isoform partners remains to be established. Using affinity purification coupled with mass spectrometry, we mapped the nuclear interactome of the FMRP isoform iso6 in U2OS. In doing so, we found FMRP nuclear interaction partners to be involved in RNA processing, pre-mRNA splicing, ribosome biogenesis, DNA replication and damage response, chromatin remodeling and chromosome segregation. By comparing interactions between nuclear iso6 and cytoplasmic iso1, we report a set of partners that bind specifically to the nuclear isoforms, mainly proteins involved in DNA-associated processes and proteasomal proteins, which is consistent with our finding that proteasome targets the nuclear FMRP iso6. The specific interactions with the nuclear isoform 6 are regulated by replication stress, while those with the cytoplasmic isoform 1 are largely insensitive to such stress, further supporting a specific role of nuclear isoforms in DNA damage response induced by replicative stress, potentially regulated by the proteasome.
Funder
Natural Sciences and Engineering Research Council of Canada Cancer Research Society
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