Stimulated PI3K-AKT signaling mediated through ligand or radiation-induced EGFR depends indirectly, but not directly, on constitutive K-Ras activity.
Previous results showed an inducible radiation sensitivity selectively observable for K-RAS-mutated cell lines as a function of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor blockade of phosphatidylinositol 3-kinase (PI3K)-AKT signaling. Therefore, the role of K-Ras activity for a direct (i.e., through activation of PI3K by K-Ras) or an indirect stimulation of PI3K-AKT signaling (through K-Ras activity-dependent EGFR ligand production) was investigated by means of small interfering RNA and inhibitor approaches as well as ELISA measurements of EGFR ligand production. K-RASmt tumor cells presented a constitutively activated extracellular signal-regulated kinase-1/2 signaling, resulting in enhanced production and secretion of the EGFR ligand amphiregulin (AREG). Medium supernatants conditioned by K-RASmt tumor cells equally efficiently stimulated EGFR signaling into the PI3K-AKT and mitogen-activated protein kinase pathways. Knocking down K-Ras expression by specific small interfering RNA markedly affected autocrine production of AREG, but not PI3K-AKT signaling, after treatment of K-RAS-mutated or wild-type cells with EGFR ligands or exposure to ionizing radiation. These results indicate that PI3K-mediated activation of AKT in K-RASmt human tumor cells as a function of EGFR ligand or radiation stimulus is independent of a direct function of K-Ras enzyme activity but depends on a K-Ras-mediated enhanced production of EGFR ligands (i.e., most likely AREG) through up-regulated extracellular signal-regulated kinase-1/2 signaling. The data provide new differential insight into the importance of K-RAS mutation in the context of PI3K-AKT-mediated radioresistance of EGFR-overexpressing or EGFR-mutated tumors.
Toulany
Mahmoud
M
Division of Radiobiology and Molecular Environmental Research, Department of Radiation Oncology, Eberhard-Karls University Tuebingen, Roentgenweg 11, 72076 Tuebingen, Germany.
Baumann
Michael
M
Rodemann
H Peter
HP
D066258:Amphiregulin; D015153:Blotting, Western; D066255:EGF Family of Proteins; D004797:Enzyme-Linked Immunosorbent Assay; D066246:ErbB Receptors / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000528:radiation effects; D011905:Genes, ras / Q000502:physiology*; D006023:Glycoproteins / Q000494:pharmacology; D006801:Humans; D047468:Immunoprecipitation; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D008024:Ligands; D009369:Neoplasms / Q000378:metabolism / Q000473:pathology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects / Q000528:radiation effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D011839:Radiation, Ionizing; D015398:Signal Transduction / Q000187:drug effects / Q000528:radiation effects; D012867:Skin / Q000166:cytology / Q000187:drug effects / Q000528:radiation effects; D014407:Tumor Cells, Cultured / Q000528:radiation effects
C587848:AREG protein, human; D066258:Amphiregulin; D066255:EGF Family of Proteins; D006023:Glycoproteins; D036341:Intercellular Signaling Peptides and Proteins; D008024:Ligands; D034741:RNA, Small Interfering; D019869:Phosphatidylinositol 3-Kinases; D066246:ErbB Receptors; C494918:AKT1 protein, human; D051057:Proto-Oncogene Proteins c-akt