title: The cyclin-dependent kinase inhibitor p27Kip1 is stabilized in G(0) by Mirk/dyrk1B kinase.
abstract: Elevated levels of the cyclin-dependent kinase (CDK) inhibitor p27 block the cell in G(0)/G(1) until mitogenic signals activate G(1) cyclins and initiate proliferation. Post-translational regulation of p27 by different phosphorylation events is critical in allowing cells to proceed through the cell cycle. We now demonstrate that the arginine-directed kinase, Mirk/dyrk1B, is maximally active in G(0) in NIH3T3 cells, when it stabilizes p27 by phosphorylating it at Ser-10. The phospho-mimetic mutant p27-S10D was more stable, and the non-phosphorylatable mutant p27-S10A was less stable than wild-type when expressed in G(0)-arrested cells. Following phosphorylation by Mirk, p27 remains a functional CDK inhibitor, capable of binding to CDK2. Mirk did not induce the translocation of p27 from the nucleus in G(0), but instead co-localized with nuclear p27. Depletion of Mirk by RNA interference decreased the phosphorylation of p27 at Ser-10 and the stability of endogenous p27. RNA(i) to Mirk increased cell entry from G(0) into G(1) as shown by increased expression of proliferating cell nuclear antigen and decreased expression of p27. These data suggest a model in which Mirk increases the amount of nuclear p27 by stabilizing it during G(0) when Mirk is most abundant. Mitogen stimulation then causes cells to enter G(1), reduces Mirk levels (Deng, X., Ewton, D., Pawlikowski, B., Maimone, M., and Friedman, E. (2003) J. Biol. Chem. 278, 41347-41354), and initiates the translocation of p27 to the cytoplasm. In addition, depletion of Mirk by RNA(i) in postmitotic C2C12 myoblasts decreased protein but not mRNA levels of p27, suggesting that stabilization of p27 by Mirk also occurs during differentiation.
authors.affiliation: Upstate Medical University, State University of New York, Syracuse, New York 13210, USA.
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Deng
authors.forename: Stephen E
authors.initials: SE
authors.lastname: Mercer
authors.forename: Sejal
authors.initials: S
authors.lastname: Shah
authors.forename: Daina Z
authors.initials: DZ
authors.lastname: Ewton
authors.forename: Eileen
authors.initials: E
authors.lastname: Friedman
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000502:physiology*; D002454:Cell Differentiation; D002455:Cell Division; D002460:Cell Line; D002478:Cells, Cultured; D050760:Cyclin-Dependent Kinase Inhibitor p27; D003513:Cycloheximide / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005982:Glutathione Transferase / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008938:Mitosis; D009154:Mutation; D041681:NIH 3T3 Cells; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D034622:RNA Interference; D011994:Recombinant Proteins / Q000737:chemistry; D016192:Resting Phase, Cell Cycle; D013997:Time Factors; D014157:Transcription Factors; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000502:physiology*; D000097603:Dyrk Kinases
chemical_list: D018797:Cell Cycle Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D003513:Cycloheximide; D004791:Enzyme Inhibitors; D005982:Glutathione Transferase; D020928:Mitogen-Activated Protein Kinases; D010744:Phosphoric Monoester Hydrolases; D018809:Proliferating Cell Nuclear Antigen; D017346:Protein Serine-Threonine Kinases; D011500:Protein Synthesis Inhibitors; D011505:Protein-Tyrosine Kinases; D011994:Recombinant Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D000097603:Dyrk Kinases; C493961:Cdkn1b protein, mouse