PKCeta associates with cyclin E/Cdk2 complex in serum-starved MCF-7 and NIH-3T3 cells.
Protein kinase C (PKC) encodes a family of enzymes implicated in cellular differentiation, growth control, and tumor promotion. However, very little is known with respect to the molecular mechanisms that link protein kinase C to cell cycle control. Here we report that PKCeta associates with the cyclin E/Cdk2 complex. This is shown for the ectopically overexpressed PKCeta in NIH-3T3 cells, the inducibly expressed PKCeta in MCF-7 cells (under control of the tetracycline-responsive promoter), and the endogenously expressed PKCeta in mouse mammary epithelial HC11 cells. Subcellular cell fractionation experiments revealed that the complex with cyclin E is formed mostly in the nuclear fractions, although in these cells PKCeta is predominantly expressed in the cytosolic fractions. The complex of PKCeta and cyclin E was studied at various phases of the cell cycle, in serum-starved quiescent cells and in cells stimulated with serum to reenter the cell cycle. Interestingly, the interaction between PKCeta and cyclin E was most prominent in serum-starved cells and was disintegrated when cells entered the cells cycle. Immunofluorescence staining demonstrated that in serum-starved cells PKCeta is concentrated at the perinuclear zone, which is also the site of its colocalization with cyclin E. Colocalization of PKCeta and cyclin E in the perinuclear region was observed in serum-starved cells, and less in proliferating cells. These experiments suggest that the interaction between PKCeta and cyclin E is carefully regulated, and is correlated with the inactivated form of the cyclin E/Cdk2 complex. Thus, our studies support an important link between PKC and cell cycle control.
Department of Microbiology and Immunology, Faculty of Health Sciences and the Cancer Research Center, Ben Gurion University, Beer Sheva 84105, Israel.
Marat
M
Shtutman
Tzippi
T
Hershko
Adva
A
Maissel
Eyal
E
Fima
Etta
E
Livneh
D016475:3T3 Cells; D000230:Adenocarcinoma; D000818:Animals; D001943:Breast Neoplasms; D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle / Q000502:physiology; D002458:Cell Fractionation; D002467:Cell Nucleus / Q000378:metabolism; D016895:Culture Media, Serum-Free; D019927:Cyclin E / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D046911:Macromolecular Substances; D051379:Mice; D011493:Protein Kinase C / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D016895:Culture Media, Serum-Free; D019927:Cyclin E; D046911:Macromolecular Substances; C081975:protein kinase C eta; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495893:Cdk2 protein, mouse; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases