Role of LXCXE motif-dependent interactions in the activity of the retinoblastoma protein.
Cell cycle control by pRb requires the integrity of the pocket domain, which is a region necessary for interactions with a variety of proteins, including E2F and LXCXE-motif containing proteins. Through knowledge of the crystal structure of pRb we have prepared a panel of pRb mutant derivatives in which a ... cluster of lysine residues that demark the LXCXE peptide binding domain were systematically mutated. One of the mutant derivatives, Rb6A, exhibits significantly reduced LXCXE-dependent interactions with HPV E7, cyclinD1 and HDAC2, but retained LXCXE-independent binding to E2F. Consistent with these results, Rb6A could down-regulate E2F-1-dependent activation of different E2F responsive promoters, but was compromised in Rb-dependent repression. Most importantly, Rb6A retained wild-type growth arrest activity, and colony forming activity similar to wild-type pRb. It is compatible with these results that directly targeting HDAC2 to E2F responsive promoters as an E2F/HDAC hybrid protein failed to effect cell cycle arrest. These results suggest that LXCXE-dependent interactions are not essential for pRb to exert growth arrest.
Mesh Terms:
Amino Acid Motifs, Amino Acid Sequence, Cell Cycle, Cell Cycle Proteins, Cell Division, Cyclin D1, DNA Mutational Analysis, DNA-Binding Proteins, Down-Regulation, E2F Transcription Factors, E2F1 Transcription Factor, Epitopes, Flow Cytometry, Glutathione Transferase, Hela Cells, Histone Deacetylase 2, Histone Deacetylases, Humans, Immunoblotting, Lysine, Models, Molecular, Molecular Sequence Data, Mutation, Peptides, Plasmids, Precipitin Tests, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, Proto-Oncogene Proteins c-myc, Recombinant Fusion Proteins, Repressor Proteins, Retinoblastoma Protein, Sequence Homology, Amino Acid, Transcription Factors, Transcription, Genetic, Transfection, Tumor Cells, Cultured
Amino Acid Motifs, Amino Acid Sequence, Cell Cycle, Cell Cycle Proteins, Cell Division, Cyclin D1, DNA Mutational Analysis, DNA-Binding Proteins, Down-Regulation, E2F Transcription Factors, E2F1 Transcription Factor, Epitopes, Flow Cytometry, Glutathione Transferase, Hela Cells, Histone Deacetylase 2, Histone Deacetylases, Humans, Immunoblotting, Lysine, Models, Molecular, Molecular Sequence Data, Mutation, Peptides, Plasmids, Precipitin Tests, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, Proto-Oncogene Proteins c-myc, Recombinant Fusion Proteins, Repressor Proteins, Retinoblastoma Protein, Sequence Homology, Amino Acid, Transcription Factors, Transcription, Genetic, Transfection, Tumor Cells, Cultured
Oncogene
Date: Sep. 27, 2001
PubMed ID: 11593423
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