RESUMO
Polycomb group (PcG) proteins form multimeric chromatin-associated protein complexes that are involved in heritable repression of gene activity. Two distinct human PcG complexes have been characterized. The EED/EZH2 PcG complex utilizes histone deacetylation to repress gene activity. The HPC/HPH PcG complex contains the HPH, RING1, BMI1, and HPC proteins. Here we show that vertebrate Polycomb homologs HPC2 and XPc2, but not M33/MPc1, interact with the histone lysine methyltransferase (HMTase) SUV39H1 both in vitro and in vivo. We further find that overexpression of SUV39H1 induces selective nuclear relocalization of HPC/HPH PcG proteins but not of the EED/EZH2 PcG proteins. This SUV39H1-dependent relocalization concentrates the HPC/HPH PcG proteins to the large pericentromeric heterochromatin domains (1q12) on human chromosome 1. Within these PcG domains we observe increased H3-K9 methylation. Finally, we show that H3-K9 HMTase activity is associated with endogenous HPC2. Our findings suggest a role for the SUV39H1 HMTase and histone H3-K9 methylation in the targeting of human HPC/HPH PcG proteins to modified chromatin structures.
Assuntos
Cromossomos/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Metiltransferases/metabolismo , Proteínas Repressoras/metabolismo , Transporte Ativo do Núcleo Celular/fisiologia , Núcleo Celular/metabolismo , Centrômero/metabolismo , Cromossomos Humanos Par 1/metabolismo , Células HeLa , Histona-Lisina N-Metiltransferase/genética , Humanos , Ligases , Substâncias Macromoleculares , Metilação , Metiltransferases/genética , Proteínas do Grupo Polycomb , Testes de Precipitina , Ligação Proteica/fisiologia , Estrutura Terciária de Proteína/fisiologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/classificação , Proteínas Repressoras/genética , Homologia de Sequência de Aminoácidos , Técnicas do Sistema de Duplo-Híbrido , Ubiquitina-Proteína LigasesRESUMO
Polycomb-group (PcG) proteins are chromatin-associated proteins that heritably repress gene activity in many organisms, including man. Two distinct human PcG complexes have been identified. The HPC/HPH PcG complex I contains the HPC, HPH, RING1, and BMI1 proteins, the EED/EZH2 PcG complex II contains the EED, EZH2, and YY1 proteins. Previously we found that the relative expression levels of proteins of the human PcG complexes I and II are severely deregulated in human tumors. These findings signify an important role for antibodies against human PcG proteins as diagnostic tools. To be able to produce standardized anti-human PcG antibodies, we developed a panel of five mouse monoclonal antibodies (MAbs) against the human PcG proteins HPC2, BMI1, RING1A, EED, and EZH2. All MAbs can be used for Western blot analysis and immunofluorescence labeling of tissue culture cells. With the exception of the MAb against HPC2, all MAbs can also be used in immunoprecipitation experiments and immunohistochemistry of human tissues. The novel MAbs are therefore valuable tools for the cell biological, biochemical, and pathological analysis of human PcG proteins.