Truncated HP1 lacking a functional chromodomain induces heterochromatinization upon in vivo targeting

Swammerdam Institute for Life Sciences, BioCentrum Amsterdam, University of Amsterdam, Kruislaan 318, 1098 SM, Amsterdam, The Netherlands.
Histochemie (Impact Factor: 3.05). 02/2006; 125(1-2):53-61. DOI: 10.1007/s00418-005-0088-7
Source: PubMed


Packaging of the eukaryotic genome into higher order chromatin structures is tightly related to gene expression. Pericentromeric heterochromatin is typified by accumulations of heterochromatin protein 1 (HP1), methylation of histone H3 at lysine 9 (MeH3K9) and global histone deacetylation. HP1 interacts with chromatin by binding to MeH3K9 through the chromodomain (CD). HP1 dimerizes with itself and binds a variety of proteins through its chromoshadow domain. We have analyzed at the single cell level whether HP1 lacking its functional CD is able to induce heterochromatinization in vivo. We used a lac-operator array-based system in mammalian cells to target EGFP-lac repressor tagged truncated HP1alpha and HP1beta to a lac operator containing gene-amplified chromosome region in living cells. After targeting truncated HP1alpha or HP1beta we observe enhanced tri-MeH3K9 and recruitment of endogenous HP1alpha and HP1beta to the chromosome region. We show that CD-less HP1alpha can induce chromatin condensation, whereas the effect of truncated HP1beta is less pronounced. Our results demonstrate that after lac repressor-mediated targeting, HP1alpha and HP1beta without a functional CD are able to induce heterochromatinization.

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Available from: Maartje C Brink
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    • "Recently, we showed that in vivo targeting of HP1␣ or HP1␤ to an amplified chromosomal region causes local chromatin condensation , enhanced trimethylated H3K9me, and recruitment of histone methyltransferases (Verschure et al., 2005). Targeting of the HP1 lacking a functional CD also caused heterochromatinization of the amplified chromosome region (Brink et al., 2006). These results show that normal binding of HP1 through its CD domain, as well as artificial binding of HP1 without a CD through lac operator-lac Repressor interaction, is sufficient to trigger heterochromatin formation. "
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    ABSTRACT: The heterochromatin protein 1 (HP1) family is thought to be an important structural component of heterochromatin. HP1 proteins bind via their chromodomain to nucleosomes methylated at lysine 9 of histone H3 (H3K9me). To investigate the role of HP1 in maintaining heterochromatin structure, we used a dominant negative approach by expressing truncated HP1alpha or HP1beta proteins lacking a functional chromodomain. Expression of these truncated HP1 proteins individually or in combination resulted in a strong reduction of the accumulation of HP1alpha, HP1beta, and HP1gamma in pericentromeric heterochromatin domains in mouse 3T3 fibroblasts. The expression levels of HP1 did not change. The apparent displacement of HP1alpha, HP1beta, and HP1gamma from pericentromeric heterochromatin did not result in visible changes in the structure of pericentromeric heterochromatin domains, as visualized by DAPI staining and immunofluorescent labeling of H3K9me. Our results show that the accumulation of HP1alpha, HP1beta, and HP1gamma at pericentromeric heterochromatin domains is not required to maintain DAPI-stained pericentromeric heterochromatin domains and the methylated state of histone H3 at lysine 9 in such heterochromatin domains.
    Full-text · Article · May 2007 · Molecular Biology of the Cell
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    • "We used the Argos system to analyze our data within the context of our research concerning chromatin structure and gene control. We focused on the effect of targeting a protein, Heterochromatin protein 1 (HP1) that is involved in gene regulation, to a defined chromosomal domain in the cell nucleus [26] [6]. We used a cell line containing a large 200 Mbp chromosome domain consisting of lac operator repeats. "
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    ABSTRACT: In this paper we propose an approach in which interactive visualization and analysis are combined with batch tools for the processing of large data collections. Large and heterogeneous data collections are difficult to analyze and pose specific problems to interactive visualization. Application of the traditional interactive processing and visualization approaches as well as batch processing encounter considerable drawbacks for such large and heterogeneous data collections due to the amount and type of data. Computing resources are not sufficient for interactive exploration of the data and automated analysis has the disadvantage that the user has only limited control and feedback on the analysis process. In our approach, an analysis procedure with features and attributes of interest for the analysis is defined interactively. This procedure is used for off-line processing of large collections of data sets. The results of the batch process along with "visual summaries" are used for further analysis. Visualization is not only used for the presentation of the result, but also as a tool to monitor the validity and quality of the operations performed during the batch process. Operations such as feature extraction and attribute calculation of the collected data sets are validated by visual inspection. This approach is illustrated by an extensive case study, in which a collection of confocal microscopy data sets is analyzed.
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    • "Hs-beta , or a CSD to reporter genes in mammalian cell culture studies caused H3K9me3, in this case by SETDB1 (BRINK et al. 2006; VERSCHURE et al. 2005). Likewise, targeting HP1 "
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    ABSTRACT: ABSTRACT Centric regions of eukaryotic genomes are packaged into heterochromatin, which possesses the ability to spread along the chromosome,and silence gene expression. The process of spreading has been challenging,to study at the molecular,level due to repetitious sequences within centric regions. An HP1 tethering system,was,developed,that generates “ectopic
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