Abstract
NIH 3T3 cells cultured in suspension fail to express cyclin A and hence cannot enter S phase and divide. We show that loss of cell adhesion to substratum abrogates cyclin A gene expression by blocking its promoter activity through the E2F site that mediates its cell cycle regulation in adherent cells. In suspended cells, G0-specific E2F complexes remain bound to the cyclin A promoter. Overexpression of cyclin D1 restores cyclin A transcription in suspended cells and rescues them from cell cycle arrest. In suspended cells, cyclin D1 and cyclin E accumulate normally upon serum stimulation, but their associated kinases remain inactive; their substrates, pRb and p107, are not hyperphosphorylated. Concomitantly, the cyclin-dependent kinase inhibitor, p27KIP1, is stabilized. Ectopic expression of p27KIP1 blocks cyclin A promoter activity through its EN binding site. These data suggest that the block to cyclin A transcription in nonadherent NIH 3T3 cells results from stabilization of p27KIP1 and subsequent inactivation of the specific E2F moiety required for its induction.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3T3 Cells
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Animals
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Base Sequence
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Carrier Proteins*
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Cell Adhesion / physiology*
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Cell Cycle Proteins*
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Cyclin D1
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Cyclin-Dependent Kinase Inhibitor p27
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Cyclins / biosynthesis
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Cyclins / genetics*
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Cyclins / physiology
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DNA-Binding Proteins*
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E2F Transcription Factors
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Gene Expression Regulation*
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Mice
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Microtubule-Associated Proteins / physiology
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Molecular Sequence Data
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Oncogene Proteins / genetics
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Oncogene Proteins / physiology
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Promoter Regions, Genetic
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Recombinant Fusion Proteins / biosynthesis
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Resting Phase, Cell Cycle / genetics
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Retinoblastoma-Binding Protein 1
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Transcription Factor DP1
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Transcription Factors / physiology*
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Transcription, Genetic*
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Tumor Suppressor Proteins*
Substances
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Arid4a protein, mouse
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Carrier Proteins
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Cdkn1b protein, mouse
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Cell Cycle Proteins
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Cyclins
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DNA-Binding Proteins
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E2F Transcription Factors
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Microtubule-Associated Proteins
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Oncogene Proteins
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Recombinant Fusion Proteins
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Retinoblastoma-Binding Protein 1
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Transcription Factor DP1
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Transcription Factors
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Tumor Suppressor Proteins
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Cyclin D1
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Cyclin-Dependent Kinase Inhibitor p27