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Pigment epithelium-derived factor (PEDF) interacts with transportin SR2, and active nuclear import is facilitated by a novel nuclear localization motif
- Source :
- PLoS ONE, Vol 6, Iss 10, p e26234 (2011), PLoS ONE
- Publication Year :
- 2011
- Publisher :
- Public Library of Science (PLoS), 2011.
-
Abstract
- PEDF (Pigment epithelium-derived factor) is a non-inhibitory member of the serpin gene family (serpinF1) that displays neurotrophic and anti-angiogenic properties. PEDF contains a secretion signal sequence, but although originally regarded as a secreted extracellular protein, endogenous PEDF is found in the cytoplasm and nucleus of several mammalian cell types. In this study we employed a yeast two-hybrid interaction trap screen to identify transportin-SR2, a member of the importin-β family of nuclear transport karyopherins, as a putative PEDF binding partner. The interaction was supported in vitro by GST-pulldown and co-immunoprecipitation. Following transfection of HEK293 cells with GFP-tagged PEDF the protein was predominantly localised to the nucleus, suggesting that active import of PEDF occurs. A motif (YxxYRVRS) shared by PEDF and the unrelated transportin-SR2 substrate, RNA binding motif protein 4b, was identified and we investigated its potential as a nuclear localization signal (NLS) sequence. Site-directed mutagenesis of this helix A motif in PEDF resulted in a GFP-tagged mutant protein being excluded from the nucleus, and mutation of two arginine residues (R67, R69) was sufficient to abolish nuclear import and PEDF interaction with transportin-SR2. These results suggest a novel NLS and mechanism for serpinF1 nuclear import, which may be critical for anti-angiogenic and neurotrophic function.
- Subjects :
- Proteomics
Models, Molecular
Amino Acid Motifs
Nuclear Localization Signals
lcsh:Medicine
Protein Synthesis
Biochemistry
Protein Structure, Secondary
Mutant protein
Molecular Cell Biology
Macromolecular Structure Analysis
Signaling in Cellular Processes
lcsh:Science
Multidisciplinary
Physics
beta Karyopherins
Cellular Structures
medicine.anatomical_structure
Protein Classes
Karyopherins
Cellular Types
Research Article
Signal Transduction
Protein Binding
Signal peptide
Protein Structure
Molecular Sequence Data
Biophysics
Active Transport, Cell Nucleus
Biology
Protein Chemistry
PEDF
Two-Hybrid System Techniques
medicine
Humans
NLS
Amino Acid Sequence
Nerve Growth Factors
Protein Interactions
Eye Proteins
Serpins
Cell Nucleus
lcsh:R
Proteins
Computational Biology
Epithelial Cells
Molecular Development
Signaling
Peptide Fragments
Signaling Networks
Cell nucleus
HEK293 Cells
Mutagenesis, Site-Directed
lcsh:Q
Nuclear transport
Nuclear localization sequence
Developmental Biology
Subjects
Details
- Language :
- English
- ISSN :
- 19326203
- Volume :
- 6
- Issue :
- 10
- Database :
- OpenAIRE
- Journal :
- PLoS ONE
- Accession number :
- edsair.doi.dedup.....b784c2426786374eb5311679699a48fc