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Dimerization of Receptor Protein-Tyrosine Phosphatase alpha in living cells
- Source :
- BMC Cell Biology, Vol 2, Iss 1, p 8 (2001), BMC Cell Biology
- Publication Year :
- 2001
- Publisher :
- BMC, 2001.
-
Abstract
- Background Dimerization is an important regulatory mechanism of single membrane-spanning receptors. For instance, activation of receptor protein-tyrosine kinases (RPTKs) involves dimerization. Structural, functional and biochemical studies suggested that the enzymatic counterparts of RPTKs, the receptor protein-tyrosine phosphatases (RPTPs), are inhibited by dimerization, but whether RPTPs actually dimerize in living cells remained to be determined. Results In order to assess RPTP dimerization, we have assayed Fluorescence Resonance Energy Transfer (FRET) between chimeric proteins of cyan- and yellow-emitting derivatives of green fluorescent protein, fused to RPTPα, using three different techniques: dual wavelength excitation, spectral imaging and fluorescence lifetime imaging. All three techniques suggested that FRET occurred between RPTPα -CFP and -YFP fusion proteins, and thus that RPTPα dimerized in living cells. RPTPα dimerization was constitutive, extensive and specific. RPTPα dimerization was consistent with cross-linking experiments, using a non-cell-permeable chemical cross-linker. Using a panel of deletion mutants, we found that the transmembrane domain was required and sufficient for dimerization. Conclusions We demonstrate here that RPTPα dimerized constitutively in living cells, which may be mediated by the transmembrane domain, providing strong support for the model that dimerization is involved in regulation of RPTPs.
- Subjects :
- Recombinant Fusion Proteins
Green Fluorescent Proteins
Phosphatase
Receptors, Cell Surface
Protein tyrosine phosphatase
Biology
Fluorescence
Green fluorescent protein
Tumor Cells, Cultured
lcsh:QH573-671
Receptor
Kinase
lcsh:Cytology
Receptor-Like Protein Tyrosine Phosphatases, Class 4
Cell Biology
Fusion protein
Protein Structure, Tertiary
Cell biology
Luminescent Proteins
Transmembrane domain
Cross-Linking Reagents
Spectrometry, Fluorescence
Förster resonance energy transfer
Energy Transfer
Microscopy, Fluorescence
Protein Tyrosine Phosphatases
Dimerization
Research Article
Subjects
Details
- Language :
- English
- ISSN :
- 14712121
- Volume :
- 2
- Issue :
- 1
- Database :
- OpenAIRE
- Journal :
- BMC Cell Biology
- Accession number :
- edsair.doi.dedup.....8de2510a82bc4535d5baf13dca20262e