title: Quantitative phosphoproteomics of murine Fmr1-KO cell lines provides new insights into FMRP-dependent signal transduction mechanisms.
abstract: Fragile X mental retardation protein (FMRP) is an RNA-binding protein that has a major effect on neuronal protein synthesis. Transcriptional silencing of the FMR1 gene leads to loss of FMRP and development of Fragile X syndrome (FXS), the most common known hereditary cause of intellectual impairment and autism. Here we utilize SILAC-based quantitative phosphoproteomics to analyze murine FMR1(-) and FMR1(+) fibroblastic cell lines derived from FMR1-KO embryos to identify proteins and phosphorylation sites dysregulated as a consequence of FMRP loss. We quantify FMRP-related changes in the levels of 5,023 proteins and 6,133 phosphorylation events and map them onto major signal transduction pathways. Our study confirms global downregulation of the MAPK/ERK pathway and decrease in phosphorylation level of ERK1/2 in the absence of FMRP, which is connected to attenuation of long-term potentiation. We detect differential expression of several key proteins from the p53 pathway, pointing to the involvement of p53 signaling in dysregulated cell cycle control in FXS. Finally, we detect differential expression and phosphorylation of proteins involved in pre-mRNA processing and nuclear transport, as well as Wnt and calcium signaling, such as PLC, PKC, NFAT, and cPLA2. We postulate that calcium homeostasis is likely affected in molecular pathogenesis of FXS.
authors.affiliation: Proteome Center Tübingen and ‡Graduate School of Cellular and Molecular Neuroscience, University of Tübingen , Österbergstrasse 3, 72074 Tübingen, Germany.
authors.forename: Katarina
authors.initials: K
authors.lastname: Matic
authors.forename: Timo
authors.initials: T
authors.lastname: Eninger
authors.forename: Barbara
authors.initials: B
authors.lastname: Bardoni
authors.forename: Laetitia
authors.initials: L
authors.lastname: Davidovic
authors.forename: Boris
authors.initials: B
authors.lastname: Macek
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002461:Cell Line, Transformed; D002853:Chromatography, Liquid; D051860:Fragile X Messenger Ribonucleoprotein 1 / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010750:Phosphoproteins / Q000378:metabolism*; D040901:Proteomics; D015398:Signal Transduction; D053719:Tandem Mass Spectrometry
chemical_list: D051860:Fragile X Messenger Ribonucleoprotein 1; D010750:Phosphoproteins; C497684:Fmr1 protein, mouse
keywords: FMR1; FMRP; MEF; autism; fragile X syndrome; phosphoproteomics