1. Oncogenic Ras blocks transforming growth factor-beta-induced cell-cycle arrest by degradation of p27 through a MEK/Erk/SKP2-dependent pathway
- Author
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Hein Schepers, Albertus T. J. Wierenga, Edo Vellenga, Bart J. L. Eggen, Faculty of Science and Engineering, Faculteit Medische Wetenschappen/UMCG, Groningen Biomolecular Sciences and Biotechnology, Celbiochemie, Stem Cell Aging Leukemia and Lymphoma, Guided Treatment in Optimal Selected Cancer Patients, Molecular Neuroscience and Ageing Research, and Restoring organ function by means of regenerative medicine
- Subjects
MAPK/ERK pathway ,Cancer Research ,DNA Mutational Analysis ,Cell Cycle Proteins ,chemistry.chemical_compound ,Mice ,Transforming Growth Factor beta ,MG132 ,RETINOIC ACID ,Extracellular Signal-Regulated MAP Kinases ,S-Phase Kinase-Associated Proteins ,GeneralLiterature_REFERENCE(e.g.,dictionaries,encyclopedias,glossaries) ,ACUTE MYELOGENOUS LEUKEMIA ,MEK inhibitor ,Cell Cycle ,TGF-BETA ,Hematology ,Flow Cytometry ,Recombinant Proteins ,Cell biology ,Ubiquitin ligase ,Electroporation ,ComputingMethodologies_DOCUMENTANDTEXTPROCESSING ,RNA Interference ,Signal transduction ,Cyclin-Dependent Kinase Inhibitor p27 ,medicine.drug ,Signal Transduction ,CYTOPLASMIC LOCALIZATION ,SIGNAL-TRANSDUCTION ,HL-60 Cells ,ACUTE MYELOID-LEUKEMIA ,Biology ,Transfection ,NUCLEAR EXPORT ,Cell Line, Tumor ,TGF beta signaling pathway ,Genetics ,medicine ,Animals ,Humans ,Molecular Biology ,Mitogen-Activated Protein Kinase Kinases ,CONSTITUTIVE PHOSPHORYLATION ,MEDIATED G(1) ARREST ,Base Sequence ,Tumor Suppressor Proteins ,Cell Biology ,Transforming growth factor beta ,DEPENDENT PHOSPHORYLATION ,Genes, ras ,chemistry ,Proteasome inhibitor ,biology.protein ,Cancer research ,ras Proteins - Abstract
Objective To examine whether oncogenic Ras affects transforming growth factor (TGF)-β–mediated cell-cycle arrest in hematopoietic cells and the downstream signal transduction pathway involved in the interference with TGF-β–induced cell-cycle arrest. Materials and Methods Two leukemic cell lines bearing N-Ras L61 mutations; HL-60 and TF-1, and the M1 cell line with wt Ras were investigated for their response to TGF-β. Signal transduction inhibitors, overexpression and RNA interference studies were performed to investigate the involvement of the various proteins. Results Although TGF-β signal transduction was not affected, G0-G1 arrest was absent in HL-60 and TF-1 cells due to the absence of p27. Overexpression of p27 restored TGF-β–induced cell-cycle arrest, as well as interfering in Ras-mediated signaling. The farnesyl transferase inhibitor L744832 and the MEK inhibitor U0126 both restored p27 levels and cell-cycle arrest in response to TGF-β. The absence of p27 protein is due to elevated levels of the ubiquitin ligase SKP2, which complexes with and targets p27 for degradation. RNA interference for SKP2 and treatment of these cells with the proteasome inhibitor MG132 restored p27 levels, corresponding with decreasing SKP2 levels after interfering in N-Ras signal transduction. P27, phosphorylated at threonine 187, is nuclear localized in N-Ras–containing cells. Mutation of this residue to alanine rendered p27 insensitive to degradation. Conclusion N-Ras L61 transformed cells lack a G0-G1 arrest upon TGF-β treatment due to absence of p27. p27 is degraded through a MapK-, and SKP2-dependent pathway. Overexpression of p27 results in restoration of cell-cycle arrest upon TGF-β treatment.
- Published
- 2005