Abstract
To develop an experimental model for E7-mediated anchorage-independent growth, we studied the ability of E7-expressing NIH 3T3 subclones to enter S phase when they were cultured in suspension. We found that expression of E7 prevents the inhibition of cyclin E-associated kinase and also triggers activation of cyclin A gene expression in suspension cells. A point mutation in the amino terminus of E7 prevented E7-driven rescue of cyclin E-associated kinase activity in suspension cells; however, cells with this mutation retained some ability to activate cyclin A gene expression and promote S-phase entry. Activation of cyclin A gene expression by E7 was correlated with an increased binding of free E2F to a regulatory element in the cyclin A promoter which mediates both repression of cyclin A upon loss of adhesion and its reactivation by E7. Surprisingly, expression of E7 led to a nuclear accumulation of one species of free E2F, namely, an E2F-4-DP-1 heterodimer, that is exclusively cytoplasmic in the absence of E7. Taken together, the data reported here indicate that several different E7-dependent changes of cellular-growth-regulating pathways can cooperate to allow adhesion-independent entry into S phase.
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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CDC2-CDC28 Kinases*
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Carrier Proteins*
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Cell Cycle Proteins / metabolism
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Cell Nucleus / metabolism
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Cyclin A / genetics*
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Cyclin E / metabolism
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Cyclin-Dependent Kinase 2
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Cyclin-Dependent Kinases / metabolism
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DNA-Binding Proteins / metabolism
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E2F Transcription Factors
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E2F4 Transcription Factor
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Humans
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Mice
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Oncogene Proteins, Viral / biosynthesis
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Oncogene Proteins, Viral / genetics
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Oncogene Proteins, Viral / metabolism*
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Papillomaviridae / genetics
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Papillomaviridae / metabolism*
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Papillomavirus E7 Proteins
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Protein Serine-Threonine Kinases / metabolism
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Retinoblastoma-Binding Protein 1
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S Phase
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Transcription Factor DP1
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Transcription Factors / biosynthesis
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Transcription Factors / genetics
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Transcription Factors / metabolism*
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Transcription, Genetic*
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Tumor Cells, Cultured
Substances
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Arid4a protein, mouse
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Carrier Proteins
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Cell Cycle Proteins
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Cyclin A
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Cyclin E
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DNA-Binding Proteins
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E2F Transcription Factors
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E2F4 Transcription Factor
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E2F4 protein, human
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E2f4 protein, mouse
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Oncogene Proteins, Viral
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Papillomavirus E7 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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oncogene protein E7, Human papillomavirus type 16
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Protein Serine-Threonine Kinases
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CDC2-CDC28 Kinases
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CDK2 protein, human
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Cdk2 protein, mouse
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Cyclin-Dependent Kinase 2
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Cyclin-Dependent Kinases