Characterization of the Akt2 Domain Essential for Binding Nuclear p21cip1 to Promote Cell Cycle Arrest during Myogenic Differentiation
| Autor: | Celine Franckhauser, Ned J.C. Lamb, Anne Fernandez, Lisa Heron-Milhavet |
|---|---|
| Přispěvatelé: | Institut de génétique humaine (IGH), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Larose, Catherine, Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Université de Montpellier (UM)-Institut National de la Santé et de la Recherche Médicale (INSERM) |
| Jazyk: | angličtina |
| Rok vydání: | 2013 |
| Předmět: |
Cyclin-Dependent Kinase Inhibitor p21
Cellular differentiation Amino Acid Motifs Blotting Western Molecular Sequence Data Cell Fluorescent Antibody Technique Sequence Homology lcsh:Medicine Peptide [SDV.GEN] Life Sciences [q-bio]/Genetics [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] Biology Muscle Development MyoD Mice 03 medical and health sciences 0302 clinical medicine [SDV.BC.BC] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] medicine Animals Humans Amino Acid Sequence Cloning Molecular lcsh:Science Peptide sequence ComputingMilieux_MISCELLANEOUS 030304 developmental biology chemistry.chemical_classification 0303 health sciences [SDV.GEN]Life Sciences [q-bio]/Genetics Multidisciplinary lcsh:R Cell Differentiation Cell Cycle Checkpoints Cell cycle Amino acid medicine.anatomical_structure Biochemistry chemistry 030220 oncology & carcinogenesis embryonic structures lcsh:Q Proto-Oncogene Proteins c-akt hormones hormone substitutes and hormone antagonists Research Article Binding domain |
| Zdroj: | PLoS ONE PLoS ONE, Public Library of Science, 2013, 8 (10), pp.e76987. ⟨10.1371/journal.pone.0076987⟩ PLoS ONE, 2013, 8 (10), pp.e76987. ⟨10.1371/journal.pone.0076987⟩ PLoS ONE, Vol 8, Iss 10, p e76987 (2013) |
| ISSN: | 1932-6203 |
| Popis: | The binding of the cdk inhibitor p21cip1 to Akt2 in the nucleus is an essential component in determining the specific role of Akt2 in the cell cycle arrest that precedes myogenic differentiation. Here, through a combination of biochemical and cell biology approaches, we have addressed the molecular basis of this binding. Using amino-terminal truncation of Akt2, we show that p21cip1 binds at the carboxy terminal of Akt2 since deletion of the first 400 amino acids did not affect the interaction between Akt2 and p21cip1. Pull down using carboxy terminal-truncated Akt2 protein revealed the importance of the region between amino acids 400 and 445 for the binding to p21cip1. Since Akt2_400–445 and Akt2_420–445 peptides could both bind p21cip1, this refines the binding domain on Akt2 between amino acids 420 and 445. In order to confirm these data in living cells, we developed a protocol to synchronize myoblasts at the cell cycle exit point when p21cip1 expression is induced by MyoD before myogenic differentiation. When a synthetic Akt2 peptide spanning the region (410–437) was microinjected in p21-expressing myoblasts, p21cip1 no longer localized exclusively in the nucleus, instead being redistributed throughout the cell, thus showing that injected peptide 410–437 acts to compete with the binding of endogenous Akt2 to p21cip1. Taken together, our data suggest that this 27 amino acid sequence on Akt2 is necessary and sufficient to bind p21cip1 both in vitro and in living cells. |
| Databáze: | OpenAIRE |
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