PKC-zeta is required for angiotensin II-induced activation of ERK and synthesis of C-FOS in MCF-7 cells.
We examined the signalling pathways responsible for the Ang II induction of growth in MCF-7 human breast cancer cells. Ang II in MCF-7 cells induced: (a) the translocation from the cytosol to membrane and nucleus of atypical protein kinase C-zeta (PKC-zeta) but not of PKC-alpha, -delta, - epsilon and -eta; (b) the expression of c-fos mRNA and protein; (c) the phosphorylation of the extracellular signal-regulated protein kinases 1 and 2 (ERK1/2). All these effects were due to the activation of the Ang II type I receptor (AT1) since they were blocked by the AT1 antagonist losartan. The Ang II-stimulated ERK1/2 phosphorylation was blocked by (a) high doses of staurosporine, inhibitor of PKC-zeta, and by a synthetic myristoylated peptide with sequences based on the endogenous PKC-zeta pseudosubstrate region (zeta-PS); (b) PD098059, a mitogen-activated protein kinase kinase inhibitor (MAPKK/MEK); and, moreover, (c) the inhibitors of phosphoinositide 3-kinases (PI3K), LY294002 and wortmannin, thus indicating that PI3K may act upstream of ERK1/2. The Ang II-evoked c-fos induction was blocked only by high doses of staurosporine and by zeta-PS whilst PD098059, LY294002 and wortmannin were ineffective, thus indicating that c-fos induction is not due to ERK1/2 activity. When the epidermal growth factor-receptor (EGFR) tyrosine kinase activity was inhibited by the use of its inhibitor AG1478, Ang II was still able to induce ERK1/2 phosphorylation and c-fos expression, therefore proving that the transactivation of EGFR was not required for these Ang II effects in MCF-7 cells. The previously reported proliferation of MCF-7 cells induced by Ang II was blocked by PD098059 and by wortmannin in a dose-dependent manner, thereby indicating that in MCF-7 cells the PI3K and ERK pathways mediate the mitogenic signalling of AT1. Our results suggest that in MCF-7 cells Ang II activates multiple signalling pathways involving PKC-zeta, PI3K and MAPK; of these pathways only PKC-zeta appears responsible for the induction of c-fos.
Muscella
Antonella
A
Laboratory of Cell Physiology, Dipartimento di Scienze e Tecnologie Biologiche e Ambientali, University of Lecce, Ecotekne, Lecce, Italy.
Greco
Simona
S
Elia
Maria Giovanna
MG
Storelli
Carlo
C
Marsigliante
Santo
S
D000804:Angiotensin II / Q000378:metabolism*; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism; D002455:Cell Division; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016762:Genes, fos / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D011945:Receptors, Angiotensin / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014407:Tumor Cells, Cultured; D000099221:Protein Kinase C zeta
D000804:Angiotensin II; D004791:Enzyme Inhibitors; D020928:Mitogen-Activated Protein Kinases; D011493:Protein Kinase C; D012333:RNA, Messenger; D011945:Receptors, Angiotensin; D000099221:Protein Kinase C zeta