Protein kinase Cepsilon is important for migration of neuroblastoma cells.
BACKGROUND
Migration is important for the metastatic capacity and thus for the malignancy of cancer cells. There is limited knowledge on regulatory factors that promote the migration of neuroblastoma cells. This study investigates the hypothesis that protein kinase C (PKC) isoforms regulate neuroblastoma cell motility.


METHODS
PKC isoforms were downregulated with siRNA or modulated with activators and inhibitors. Migration was analyzed with scratch and transwell assays. Protein phosphorylation and expression levels were measured with Western blot.


RESULTS
Stimulation with 12-O-tetradecanoylphorbol-13-acetate (TPA) induced migration of SK-N-BE(2)C neuroblastoma cells. Treatment with the general protein kinase C (PKC) inhibitor GF109203X and the inhibitor of classical isoforms Gö6976 inhibited migration while an inhibitor of PKCbeta isoforms did not have an effect. Downregulation of PKCepsilon, but not of PKCalpha or PKCdelta, with siRNA led to a suppression of both basal and TPA-stimulated migration. Experiments using PD98059 and LY294002, inhibitors of the Erk and phosphatidylinositol 3-kinase (PI3K) pathways, respectively, showed that PI3K is not necessary for TPA-induced migration. The Erk pathway might be involved in TPA-induced migration but not in migration driven by PKCepsilon. TPA induced phosphorylation of the PKC substrate myristoylated alanine-rich C kinase substrate (MARCKS) which was suppressed by the PKC inhibitors. Treatment with siRNA oligonucleotides against different PKC isoforms before stimulation with TPA did not influence the phosphorylation of MARCKS.


CONCLUSIONS
PKCepsilon is important for migration of SK-N-BE(2)C neuroblastoma cells. Neither the Erk pathway nor MARCKS are critical downstream targets of PKCepsilon but they may be involved in TPA-mediated migration.
Stensman
Helena
H
Lund University, Center for Molecular Pathology, Dept of Laboratory Medicine, Malmö University Hospital, Malmö, Sweden. helena.stensman@gmail.com
Larsson
Christer
C
D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology*; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D020935:MAP Kinase Signaling System; D008565:Membrane Proteins / Q000378:metabolism; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D009447:Neuroblastoma / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D051744:Protein Kinase C-epsilon / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D014162:Transfection
D047908:Intracellular Signaling Peptides and Proteins; C000619710:MARCKS protein, human; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D019869:Phosphatidylinositol 3-Kinases; D051744:Protein Kinase C-epsilon; D048049:Extracellular Signal-Regulated MAP Kinases