Abstract
The Polycomb repressive complex 1 (PRC1) mediates gene silencing, in part by monoubiquitination of histone H2A on lysine 119 (uH2A). Bmi1 and Ring1b are critical components of PRC1 that heterodimerize via their N-terminal RING domains to form an active E3 ubiquitin ligase. We have determined the crystal structure of a complex between the Bmi1/Ring1b RING-RING heterodimer and the E2 enzyme UbcH5c and find that UbcH5c interacts with Ring1b only, in a manner fairly typical of E2-E3 interactions. However, we further show that the Bmi1/Ring1b RING domains bind directly to duplex DNA through a basic surface patch unique to the Bmi1/Ring1b RING-RING dimer. Mutation of residues on this interaction surface leads to a loss of H2A ubiquitination activity. Computational modelling of the interface between Bmi1/Ring1b-UbcH5c and the nucleosome suggests that Bmi1/Ring1b interacts with both nucleosomal DNA and an acidic patch on histone H4 to achieve specific monoubiquitination of H2A. Our results point to a novel mechanism of substrate recognition, and control of product formation, by Bmi1/Ring1b.
MeSH terms
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Binding Sites
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Crystallography, X-Ray
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DNA / metabolism
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism*
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Histones / metabolism
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Humans
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Models, Molecular
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Mutagenesis, Site-Directed
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Nuclear Proteins / chemistry
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism*
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Nucleosomes / metabolism*
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Polycomb Repressive Complex 1
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Protein Binding
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Protein Conformation
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Proto-Oncogene Proteins / chemistry
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Proto-Oncogene Proteins / genetics
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Proto-Oncogene Proteins / metabolism*
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / metabolism
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Repressor Proteins / chemistry
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Repressor Proteins / genetics
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Repressor Proteins / metabolism*
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Sodium Chloride / pharmacology
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Ubiquitin-Conjugating Enzymes / chemistry
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Ubiquitin-Conjugating Enzymes / genetics
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Ubiquitin-Conjugating Enzymes / metabolism*
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Ubiquitin-Protein Ligases / chemistry
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Ubiquitin-Protein Ligases / genetics
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Ubiquitin-Protein Ligases / metabolism*
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Ubiquitination
Substances
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BMI1 protein, human
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DNA-Binding Proteins
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Histones
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Nuclear Proteins
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Nucleosomes
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Proto-Oncogene Proteins
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Recombinant Fusion Proteins
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Repressor Proteins
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Sodium Chloride
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DNA
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UBE2D3 protein, human
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Ubiquitin-Conjugating Enzymes
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Polycomb Repressive Complex 1
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RNF2 protein, human
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Ubiquitin-Protein Ligases