Isolation of an FMRP-Associated Messenger Ribonucleoprotein Particle and Identification of Nucleolin and the Fragile X-Related Proteins as Components of the Complex
Open accessAbstract
The loss of FMR1 expression due to trinucleotide repeat expansion leads to fragile X syndrome, a cause of mental retardation. The encoded protein, FMRP, is a member of a gene family that also contains the fragile X-related proteins, FXR1P and FXR2P. FMRP has been shown to be a nucleocytoplasmic shuttling protein that selectively binds a subset of mRNAs, forms messenger ribonucleoprotein (mRNP) complexes, and associates with translating ribosomes. Here we describe a cell culture system from which we can isolate epitope-tagged FMRP along with mRNA, including its own message, and at least six other proteins. We identify two of these proteins as FXR1P and FXR2P by using specific antisera and identify a third protein as nucleolin by using mass spectrometry. The presence of nucleolin is confirmed by both reactivity with a specific antiserum as well as reverse coimmunoprecipitation where antinucleolin antiserum immunoprecipitates endogenous FMRP from both cultured cells and mouse brain. The identification of nucleolin, a known component of other mRNPs, adds a new dimension to the analysis of FMRP function, and the approach described should also allow the identification of the remaining unknown proteins of this FMRP-associated mRNP as well as the other bound mRNAs.
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Nucleolin interacts with FMRP to form multiprotein complexes.
“Since nucleolin interacts with FMRP to form multiprotein complexes [38], we would expect the 29-base element to be protected from T1 ribonuclease digestion of anti-FMRP IPs, as shown in Figure 7A (nt 2318–2416).”
From FMRP Mediates mGluR5-Dependent Translation of Amyloid Precursor Protein (2007) · cites this paper as [38]
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“Since nucleolin interacts with FMRP to form multiprotein complexes [38], we would expect the 29-base element to be protected from T1 ribonuclease digestion of anti-FMRP IPs, as shown in Figure 7A (nt 2318–2416).”
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- Authors
- Ceman, S., Brown, V., & Warren, S. T.
- Affiliations
- Howard Hughes Medical Institute (US)
- Venue
- Molecular and Cellular Biology
- Publisher
- Informa UK Limited
- Published
- 1999-12
- Volume
- 19
- Issue
- 12
- Pages
- 7925-7932
- ISSN
- 0270-7306 (ISSN-L)
- 1098-5549
- PMC
- PMC84877
- OpenAlex
- W2114955795
- Cited by
- 181
- References
- 1
- Citations by year
- 2025: 1 · 2023: 1 · 2021: 2 · 2020: 3 · 2019: 1 · 2018: 4 · 2016: 4 · 2015: 3 · 2014: 10 · 2013: 3 · 2012: 6
- FWCI
- 7.77
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Molecular and Cellular Biology (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Howard Hughes Medical Institute
- Eunice Kennedy Shriver National Institute of Child Health and Human Development (R37 HD20521, P01 HD35576, R37HD020521, P01HD035576)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genetics and Neurodevelopmental Disorders
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Keywords
- FMRP, fragile X syndrome, messenger ribonucleoprotein, nucleolin, FXR1P, trinucleotide repeat expansion, mRNA binding, protein complex isolation, nucleocytoplasmic shuttling, mouse brain
- MeSH terms
- Nucleolin, Animals, Cell Line, Fragile X Syndrome, Nerve Tissue Proteins, Oligopeptides, Peptides, Phosphoproteins, Precipitin Tests, Recombinant Proteins, Ribonucleases, Ribonucleoproteins, RNA, Messenger, Gene Expression, RNA-Binding Proteins, Mice, Knockout, Mice, Fragile X Messenger Ribonucleoprotein 1, Fragile X Mental Retardation Protein
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