Subcellular fractionation and localization studies reveal a direct interaction of the fragile X mental retardation protein (FMRP) with nucleolin

PLoS One. 2014 Mar 21;9(3):e91465. doi: 10.1371/journal.pone.0091465. eCollection 2014.

Abstract

Fragile X mental Retardation Protein (FMRP) is a well-known regulator of local translation of its mRNA targets in neurons. However, despite its ubiquitous expression, the role of FMRP remains ill-defined in other cell types. In this study we investigated the subcellular distribution of FMRP and its protein complexes in HeLa cells using confocal imaging as well as detergent-free fractionation and size exclusion protocols. We found FMRP localized exclusively to solid compartments, including cytosolic heavy and light membranes, mitochondria, nuclear membrane and nucleoli. Interestingly, FMRP was associated with nucleolin in both a high molecular weight ribosomal and translation-associated complex (≥6 MDa) in the cytosol, and a low molecular weight complex (∼200 kDa) in the nucleoli. Consistently, we identified two functional nucleolar localization signals (NoLSs) in FMRP that are responsible for a strong nucleolar colocalization of the C-terminus of FMRP with nucleolin, and a direct interaction of the N-terminus of FMRP with the arginine-glycine-glycine (RGG) domain of nucleolin. Taken together, we propose a novel mechanism by which a transient nucleolar localization of FMRP underlies a strong nucleocytoplasmic translocation, most likely in a complex with nucleolin and possibly ribosomes, in order to regulate translation of its target mRNAs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Fractionation
  • Fragile X Mental Retardation Protein / analysis
  • Fragile X Mental Retardation Protein / metabolism*
  • Gene Expression Regulation
  • HeLa Cells
  • Humans
  • Nuclear Localization Signals
  • Phosphoproteins / analysis
  • Phosphoproteins / metabolism*
  • Protein Transport
  • RNA, Messenger / metabolism
  • RNA-Binding Proteins / analysis
  • RNA-Binding Proteins / metabolism*

Substances

  • Nuclear Localization Signals
  • Phosphoproteins
  • RNA, Messenger
  • RNA-Binding Proteins
  • nucleolin
  • Fragile X Mental Retardation Protein

Grant support

This work was supported in part by the German Academic Exchange Service (DAAD), the International Graduate School of Protein Science and Technology (iGRASP) and by the Strategic Research Fund (SFF) of the Heinrich-Heine University Düsseldorf (Grant F-2012/79-6). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.