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The Soluble Heparin-Binding EGF-Like Growth Factor Stimulates EGF Receptor Trafficking to the Nucleus.

Korotkevych NV, Labyntsev AJ, Kolybo DV, Komisarenko SV - PLoS ONE (2015)

Bottom Line: Soluble form of heparin-binding EGF-like growth factor (sHB-EGF) is one of the ligands for EGFR in many cell types; however, there is no evidence for the ability of sHB-EGF to induce EGFR nuclear importation.Here, we demonstrated that treatment of A431 cells with sHB-EGF resulted in nuclear localization of EGFR and such translocation occurs via retrograde pathway.The obtained data suggest that sHB-EGF acts similarly to other EGFR ligands and is capable to induce EGFR nuclear translocation as a part of ligand-receptor complex in a tyrosine phosphorylation-dependent manner.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology Department, Palladin Institute of Biochemistry of the National Academy of Sciences of Ukraine, Kyiv, Ukraine.

ABSTRACT
Most ligands of epidermal growth factor receptor (EGFR) have the ability to induce EGFR translocation into the nucleus, where EGFR acts as an important transcriptional regulator. Soluble form of heparin-binding EGF-like growth factor (sHB-EGF) is one of the ligands for EGFR in many cell types; however, there is no evidence for the ability of sHB-EGF to induce EGFR nuclear importation. Here, we demonstrated that treatment of A431 cells with sHB-EGF resulted in nuclear localization of EGFR and such translocation occurs via retrograde pathway. It was shown by confocal microscopy and co-immunoprecipitation assay that the translocation complex consisted of both ligand and receptor. The chromatin immunoprecipitation assay showed the association of sHB-EGF-EGFR complex with promoter region of cyclin D1 in the cell nucleus and this association was prevented by application of EGFR kinase inhibitor AG-1478. The obtained data suggest that sHB-EGF acts similarly to other EGFR ligands and is capable to induce EGFR nuclear translocation as a part of ligand-receptor complex in a tyrosine phosphorylation-dependent manner.

No MeSH data available.


EGFR interaction with sHB-EGF and subcellular localization of their complex.Z-stack confocal image. A431 cells transfected with pEGFR-EYFP were stimulated with mCherry-sHB-EGF for 30 min. (mCherry-sHB-EGF (Red); EGFR-EYFP (green), sHB-EGF/EGFR ligand-receptor complex (yellow)). White scale bar corresponding to 10 um.
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pone.0127887.g003: EGFR interaction with sHB-EGF and subcellular localization of their complex.Z-stack confocal image. A431 cells transfected with pEGFR-EYFP were stimulated with mCherry-sHB-EGF for 30 min. (mCherry-sHB-EGF (Red); EGFR-EYFP (green), sHB-EGF/EGFR ligand-receptor complex (yellow)). White scale bar corresponding to 10 um.

Mentions: To investigate the ability of HB-EGF to form complexes with EGFR during its intracellular trafficking to the nuclear envelope we used fluorescent analogue of sHB-EGF labeled with mCherry (mCherry-sHB-EGF). A431 cells were transfected with plasmid pEGFR-EYFP and then were treated with mCherry-sHB-EGF. Hoechst 33342 was used to localize the nuclei. The ligand-receptor complex (yellow signal) was found in the perinuclear region as an overlap of green (EYFP) and red (mCherry) signals in Z-stack images (Fig 3). This data suggested that sHB-EGF interacts with EGFR during its translocation to the nuclear envelope.


The Soluble Heparin-Binding EGF-Like Growth Factor Stimulates EGF Receptor Trafficking to the Nucleus.

Korotkevych NV, Labyntsev AJ, Kolybo DV, Komisarenko SV - PLoS ONE (2015)

EGFR interaction with sHB-EGF and subcellular localization of their complex.Z-stack confocal image. A431 cells transfected with pEGFR-EYFP were stimulated with mCherry-sHB-EGF for 30 min. (mCherry-sHB-EGF (Red); EGFR-EYFP (green), sHB-EGF/EGFR ligand-receptor complex (yellow)). White scale bar corresponding to 10 um.
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Related In: Results  -  Collection

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pone.0127887.g003: EGFR interaction with sHB-EGF and subcellular localization of their complex.Z-stack confocal image. A431 cells transfected with pEGFR-EYFP were stimulated with mCherry-sHB-EGF for 30 min. (mCherry-sHB-EGF (Red); EGFR-EYFP (green), sHB-EGF/EGFR ligand-receptor complex (yellow)). White scale bar corresponding to 10 um.
Mentions: To investigate the ability of HB-EGF to form complexes with EGFR during its intracellular trafficking to the nuclear envelope we used fluorescent analogue of sHB-EGF labeled with mCherry (mCherry-sHB-EGF). A431 cells were transfected with plasmid pEGFR-EYFP and then were treated with mCherry-sHB-EGF. Hoechst 33342 was used to localize the nuclei. The ligand-receptor complex (yellow signal) was found in the perinuclear region as an overlap of green (EYFP) and red (mCherry) signals in Z-stack images (Fig 3). This data suggested that sHB-EGF interacts with EGFR during its translocation to the nuclear envelope.

Bottom Line: Soluble form of heparin-binding EGF-like growth factor (sHB-EGF) is one of the ligands for EGFR in many cell types; however, there is no evidence for the ability of sHB-EGF to induce EGFR nuclear importation.Here, we demonstrated that treatment of A431 cells with sHB-EGF resulted in nuclear localization of EGFR and such translocation occurs via retrograde pathway.The obtained data suggest that sHB-EGF acts similarly to other EGFR ligands and is capable to induce EGFR nuclear translocation as a part of ligand-receptor complex in a tyrosine phosphorylation-dependent manner.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology Department, Palladin Institute of Biochemistry of the National Academy of Sciences of Ukraine, Kyiv, Ukraine.

ABSTRACT
Most ligands of epidermal growth factor receptor (EGFR) have the ability to induce EGFR translocation into the nucleus, where EGFR acts as an important transcriptional regulator. Soluble form of heparin-binding EGF-like growth factor (sHB-EGF) is one of the ligands for EGFR in many cell types; however, there is no evidence for the ability of sHB-EGF to induce EGFR nuclear importation. Here, we demonstrated that treatment of A431 cells with sHB-EGF resulted in nuclear localization of EGFR and such translocation occurs via retrograde pathway. It was shown by confocal microscopy and co-immunoprecipitation assay that the translocation complex consisted of both ligand and receptor. The chromatin immunoprecipitation assay showed the association of sHB-EGF-EGFR complex with promoter region of cyclin D1 in the cell nucleus and this association was prevented by application of EGFR kinase inhibitor AG-1478. The obtained data suggest that sHB-EGF acts similarly to other EGFR ligands and is capable to induce EGFR nuclear translocation as a part of ligand-receptor complex in a tyrosine phosphorylation-dependent manner.

No MeSH data available.