Radiation-induced caveolin-1 associated EGFR internalization is linked with nuclear EGFR transport and activation of DNA-PK.
BACKGROUND
To elucidate the role of src kinase in caveolin-1 driven internalization and nuclear transport of EGFR linked to regulation of DNA-repair in irradiated cells.


RESULTS
Ionizing radiation resulted in src kinase stabilization, activation and subsequent src mediated caveolin-1 Y14- and EGFR Y845-phosphorylations. Both phosphorylations were radiation specific and could not be observed after treatment with EGF. Inhibition of EGFR by the antibody Erbitux resulted in a strong accumulation of caveolin/EGFR complexes within the cytoplasm, which could not be further increased by irradiation. Radiation-induced caveolin-1- and EGFR-phosphorylations were associated with nuclear EGFR transport and activation of DNA-PK, as detected by phosphorylation at T2609. Blockage of src activity by the specific inhibitor PP2, decreased nuclear transport of EGFR and inhibited caveolin-1- and DNA-PK-phosphorylation. Knockdown of src by specific siRNA blocked EGFR phosphorylation at Y845, phosphorylation of caveolin-1 at Y14 and abolished EGFR transport into the nucleus and phosphorylation of DNA-PK. Consequently, both knockdown of src by specific siRNA and also inhibition of src activity by PP2 resulted in an enhanced residual DNA-damage as quantified 24 h after irradiation and increased radiosensitivity.


CONCLUSIONS
Src kinase activation following irradiation triggered caveolin-1 dependent EGFR internalization into caveolae. Subsequently EGFR shuttled into the nucleus. As a consequence, inhibition of internalization and nuclear transport of EGFR blocked radiation-induced phosphorylation of DNA-PK and hampered repair of radiation-induced double strand breaks.
Dittmann
Klaus
K
Division of Radiobiology and Environmental Research, Department of Radiation Oncology University of Tübingen, Röntgenweg 11, 72076 Tübingen, Germany. klaus.dittmann@uni-tuebingen.de
Mayer
Claus
C
Kehlbach
Rainer
R
Rodemann
H Peter
HP
D021581:Active Transport, Cell Nucleus / Q000528:radiation effects; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D061067:Antibodies, Monoclonal, Humanized; D051242:Caveolin 1 / Q000378:metabolism*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism*; D000068818:Cetuximab; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D051747:DNA-Activated Protein Kinase / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism*; D006801:Humans; D010766:Phosphorylation / Q000528:radiation effects; D034741:RNA, Small Interfering / Q000378:metabolism; D014162:Transfection; D019061:src-Family Kinases / Q000378:metabolism
D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D051242:Caveolin 1; D034741:RNA, Small Interfering; D066246:ErbB Receptors; D019061:src-Family Kinases; D051747:DNA-Activated Protein Kinase; D000068818:Cetuximab
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