Atypical PKC-zeta and PKC-iota mediate opposing effects on MCF-7 Na+/K+ATPase activity.
We demonstrated previously that in serum-starved MCF-7 breast cancer cell line, Ang II increased Na+/K+ATPase activity and activated the protein kinase C zeta (PKC-zeta) (Muscella et al., 2002 J Endocrinol 173:315-323; 2003 J Cell Physiol 197:61-68.). The aim of the present study was to investigate the modulation of the activity of the Na+/K+ATPase by PKC-zeta in MCF-7 cells. Here, using serum-starved MCF-7 cells, we have demonstrated that the effect of Ang II on the Na+/K+ATPase activity was inhibited by a synthetic myristoylated peptide with sequences based on the endogenous PKC-zeta pseudosubstrate region (zeta-PS) and by high doses of GF109203X, inhibitor of PKCs. When MCF-7 cells, grown in 10% fetal bovine serum (FBS), were stimulated with Ang II a dose- and time-dependent inhibition of the Na+/K+ATPase activity was obtained. Under this growth condition we found that mRNAs for AT1, AT2, and for Na+/K+ATPase alpha1 and alpha3 subunits were unchanged; besides both the activity of the Na+/K+ATPase and the level of PKC-zeta also were unaffected by the serum. The atypical PKC-iota level (present in very low abundance in serum-starved MCF-7) was increased and Ang II provoked its translocation from the cytosol to plasma membrane. PKC-zeta was localized to the membrane, and upon Ang II treatment its cellular localization did not change. The Ang II-mediated decrease of the Na+/K+ATPase activity was inhibited by high doses of GF109203X but not by zeta-PS, thus indicating that such effect was not due to PKC-zeta activity. The treatment of cells with PKC-iota antisense oligodeoxynucleotides inhibited the effects of Ang II on the Na+/K+ATPase activity. Additionally, the effect of Ang II on Na+/K+ATPase activity was also blocked by the phosphatidylinositol 3-kinase (PI3K) inhibitors, wortmannin and LY294002, and by the actin depolymerizing agents, cytochalasin D. In conclusion, in MCF-7 cells Ang II modulates the Na+/K+ATPase activity by both atypical PKC-zeta/-iota. The effects of Ang II are opposite depending upon the presence of the serum-sensitive PKC-iota, with the inhibitory effect possibly due to the redistribution of sodium pump from plasma membrane to the inactive intracellular pool.
Muscella
Antonella
A
Dipartimento di Scienze e Tecnologie Biologiche e Ambientali, Universitá degli Studi di Lecce, Ecotekne, Via Provinciale per Monteroni, Lecce, Italy.
Storelli
Carlo
C
Marsigliante
Santo
S
D000730:Androstadienes / Q000494:pharmacology; D000804:Angiotensin II / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000473:pathology; D002417:Cattle / Q000097:blood / Q000196:embryology; D045744:Cell Line, Tumor; D002867:Chromones / Q000494:pharmacology; D016895:Culture Media, Serum-Free; D015638:Cytochalasin D / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008301:Maleimides / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000077191:Wortmannin; D000099293:Protein Kinase C-lambda; D000099221:Protein Kinase C zeta
D000730:Androstadienes; D000804:Angiotensin II; D002867:Chromones; D016895:Culture Media, Serum-Free; D015638:Cytochalasin D; D004791:Enzyme Inhibitors; D007211:Indoles; D007527:Isoenzymes; D008301:Maleimides; D009025:Morpholines; D020319:Oligodeoxyribonucleotides, Antisense; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C; D012333:RNA, Messenger; D000254:Sodium-Potassium-Exchanging ATPase; D000077191:Wortmannin; D000099293:Protein Kinase C-lambda; D000099221:Protein Kinase C zeta; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C070515:bisindolylmaleimide I