Methylmercury inhibits TrkA signaling through the ERK1/2 cascade after NGF stimulation of PC12 cells.
Using PC12 cells as a model of neuronal differentiation, we have shown that acute exposure to methylmercury (CH3Hg) inhibits nerve growth factor (NGF)-induced activation of TrkA. In the present study, we examined the effects of CH3Hg on pathways activated by NGF. NGF-induced phosphorylation of ERK1/2 in PC12 cells was time-dependent. Concurrent exposure to CH3Hg and NGF for 2.5 min resulted in a concentration-dependent inhibition of ERK1/2 phosphorylation (EC50 = 0.018 microM). However, NGF-stimulated ERK1/2 phosphorylation was not altered after 5 min of exposure to CH3Hg. In vitro studies revealed that CH3Hg did not directly inhibit the ERK kinase MEK. As reported in other neuronal tissue, CH3Hg can inhibit PKC activity in vitro. Incubation of PC12 cell lysates with CH3Hg produced a concentration-dependent inhibition of PKC activity that was significant at 0.3-10 microM. Further studies using recombinant enzymes examined the effect of CH3Hg on PKC isoforms expressed in PC12 cells. CH3Hg inhibited PKCdelta, and zeta activity in a concentration-dependent manner at higher concentrations (3-10 microM), while a significant increase in PKCalpha activity was observed at lower concentrations (0.1 microM). However, CH3Hg had no affect on NGF-induced PKC activity in intact cells. These results show that CH3Hg inhibition of NGF-stimulated TrkA activation in PC12 cells decreases downstream signaling through the Raf/MEK/ERK cascade. In intact cells PKC does not appear to be a primary target for CH3Hg.
Parran
Damani K
DK
Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Barone
Stanley
S
Mundy
William R
WR
D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002352:Carrier Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D006801:Humans; D048728:MAP Kinase Kinase Kinase 1; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D008767:Methylmercury Compounds / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000502:physiology*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology*; D020932:Nerve Growth Factor / Q000494:pharmacology*; D016716:PC12 Cells; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D020917:Receptor, trkA; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors
D002352:Carrier Proteins; D008565:Membrane Proteins; D008767:Methylmercury Compounds; D020932:Nerve Growth Factor; D020917:Receptor, trkA; D011493:Protein Kinase C; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048728:MAP Kinase Kinase Kinase 1; D020930:MAP Kinase Kinase Kinases; C482576:MAP3K1 protein, human