title: Lead acetate induces EGFR activation upstream of SFK and PKCalpha linkage to the Ras/Raf-1/ERK signaling.
abstract: Lead acetate (Pb), a probable human carcinogen, can activate protein kinase C (PKC) upstream of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Yet, it remains unclear whether Pb activation of PKC --> ERK1/2 involves receptor/non-receptor tyrosine kinases and the Ras signaling transducer. Here we demonstrate a novel mechanism elicited by Pb for transmitting ERK1/2 signaling in CL3 human non-small-cell lung adenocarcinoma cells. Pb induction of higher steady-state levels of Ras-GTP was essential for increasing phospho-Raf-1(S338) and phospho-ERK1/2. Pre-treatment of the cells with a conventional PKC inhibitor Gö6976 or depleting PKCalpha using specific small interfering RNA blocked Pb induction of Ras-GTP. Pb also activated cellular tyrosine kinases. Specific pharmacological inhibitors, PD153035 for epidermal growth factor receptor (EGFR) and SU6656 for Src family tyrosine kinases (SFK), but not AG1296 for platelet-derived growth factor receptor, could suppress the Pb-induced tyrosine kinases, PKCalpha, Ras-GTP, phospho-Raf-1(S338) and phospho-ERK1/2. Furthermore, phosphorylation of tyrosines on the EGFR multiple autophosphorylation sites and the conserved SFK autophosphorylation site occurred during exposure of cells to Pb for 1-5 min and 5-30 min, respectively. Intriguingly, Pb activation of EGFR required the intrinsic kinase activity but not dimerization of the receptor. Inhibition of SFK or PKCalpha activities did not affect EGFR phosphorylation, while knockdown of EGFR blocked SFK phosphorylation and PKCalpha activation following Pb. Together, these results indicate that immediate activation of EGFR in response to Pb is obligatory for activation of SFK and PKCalpha and subsequent the Ras-Raf-1-MKK1/2-ERK1/2 signaling cascade.
authors.affiliation: Molecular Carcinogenesis Laboratory, Institute of Biotechnology & Department of Life Sciences, National Tsing Hua University, Hsinchu 30013, Taiwan.
authors.forename: Chun-Yu
authors.initials: CY
authors.lastname: Wang
authors.forename: Yi-Ting
authors.initials: YT
authors.lastname: Wang
authors.forename: Der-Wan
authors.initials: DW
authors.lastname: Tzeng
authors.forename: Jia-Ling
authors.initials: JL
authors.lastname: Yang
mesh_terms: D015153:Blotting, Western; D002087:Butyrates / Q000494:pharmacology; D045744:Cell Line, Tumor; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000187:drug effects*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics*; D011905:Genes, ras / Q000235:genetics*; D006801:Humans; D047468:Immunoprecipitation; D007211:Indoles / Q000494:pharmacology; D009942:Organometallic Compounds / Q000633:toxicity*; D010636:Phenols / Q000494:pharmacology; D010766:Phosphorylation; D051571:Protein Kinase C-alpha / Q000235:genetics*; D011799:Quinazolines / Q000494:pharmacology; D017479:Receptors, Platelet-Derived Growth Factor / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology; D014162:Transfection; D048490:raf Kinases / Q000235:genetics*; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000235:genetics*
chemical_list: D002087:Butyrates; D007211:Indoles; D009942:Organometallic Compounds; D010636:Phenols; D011799:Quinazolines; C416927:SU 6656; D013449:Sulfonamides; C487958:elsibucol; D066246:ErbB Receptors; D017479:Receptors, Platelet-Derived Growth Factor; D019061:src-Family Kinases; D048490:raf Kinases; D051571:Protein Kinase C-alpha; D048049:Extracellular Signal-Regulated MAP Kinases; C008261:lead acetate; C088860:4-((3-bromophenyl)amino)-6,7-dimethoxyquinazoline