Adhesion-regulated G1 cell cycle arrest in epithelial cells requires the downregulation of c-Myc.
Adhesion to the extracellular matrix is required for the expression and activation of the cyclin-cyclin-dependent kinase (CDK) complexes, and for G1 phase progression of non-transformed cells. However, in non-adherent cells no molecular mechanism has yet been proposed for the cell adhesion-dependent up-regulation of the p27 cyclin-dependent kinase inhibitor (CKI), and the associated inhibition of cyclin E-CDK2. We now show that in epithelial cells the expression of c-Myc is tightly regulated by cell-substrate adhesion. When deprived of adhesion, two independently derived mammary epithelial cell lines, 184A1N4 and MCF-10A, rapidly decrease their level of c-Myc mRNA and protein. This decrease in levels of c-Myc correlates with G1 phase arrest, as indicated by hypophosphorylation of pRb and inhibition of the activity of the cyclin E-CDK2 complex. In 184A1N4 cells, cell-substrate adhesion is required for the suppression of p27, and induction of cyclin E, E2F-1, but not cyclins D1 and D3. Enforced expression of c-Myc in non-adherent 184A1N4 and MCF-10A cells reverses the adhesion-dependent inhibition of cell cycle progression. Restoration of c-Myc in non-adherent cells induces the expression of E2F-1, and hyperphosphorylation of pRb in response to EGF treatment. In addition, expression of c-Myc results in the anchorage-independent activation of the CDK2 complex, the associated upregulation of cyclin E, and the destabilization and degradation of p27 by the ubiquitin-proteasome pathway. Our study thus suggests that c-Myc is the link between cell adhesion and the regulation of p27 and cyclin E-CDK2. Furthermore, we describe a role for c-Myc in adhesion-mediated regulation of E2F-1.
Benaud
C M
CM
Department of Cell Biology, Vincent T Lombardi Cancer Center, Georgetown University Medical Center, 3970 Reservoir Road NW, Washington, DC 20007, USA.
Dickson
R B
RB
D001940:Breast / Q000166:cytology; D042846:CDC2-CDC28 Kinases; D002448:Cell Adhesion / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D019927:Cyclin E / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004268:DNA-Binding Proteins; D003609:Dactinomycin / Q000494:pharmacology; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D004815:Epidermal Growth Factor / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology*; D005109:Extracellular Matrix / Q000502:physiology; D005260:Female; D016193:G1 Phase / Q000502:physiology*; D016259:Genes, myc; D006801:Humans; D009097:Multienzyme Complexes / Q000378:metabolism; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000502:physiology; D016160:Retinoblastoma Protein / Q000378:metabolism; D015398:Signal Transduction; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D025521:Tumor Suppressor Proteins; D014452:Ubiquitins / Q000378:metabolism
D018797:Cell Cycle Proteins; D019927:Cyclin E; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D009097:Multienzyme Complexes; D019384:Nucleic Acid Synthesis Inhibitors; D011500:Protein Synthesis Inhibitors; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D016160:Retinoblastoma Protein; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D014452:Ubiquitins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D003609:Dactinomycin; D004815:Epidermal Growth Factor; D003513:Cycloheximide; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495900:CDK4 protein, human; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex