Heterochromatin protein 1 distribution during development and during the cell cycle in Drosophila embryos

Heterochromatin protein 1 (HP1) was initially discovered as a protein that is associated with the heterochromatin at the chromocenter of polytene chromosomes in Drosophila larval salivary glands. In this paper we investigate the localization of heterochromatin protein 1 in the diploid nuclei of Drosophila embryos. We focus on its association with the interphase heterochromatin in fixed embryos before and during cycle 14, the developmental time at which heterochromatin becomes most conspicuous, and also follow its localization during mitosis. The GAGA transcription factor was recently shown to be localized at sequences within alpha-heterochromatin in pre-cycle 14 embryos, and an antibody against this protein serves as a convenient marker for these sequences. We find an enrichment of heterochromatin protein 1 in the intensely DAPI-staining regions near the apical surface of nuclear cycle 10 embryos. At this stage GAGA factor is localized into punctate structures in this same region. This enrichment for HP1 is markedly increased during nuclear cycle 14. Surprisingly, whereas GAGA factor retains its association with the heterochromatin throughout the cell cycle, a significant fraction of HP1 is dispersed throughout the spindle around the segregating chromosomes during mitosis. This dispersed pool of heterochromatin protein 1 was observed during mitosis in both early and late Drosophila embryos and in an analysis of a bacterially produced 6x histidine-heterochromatin protein 1 fusion protein injected into living Drosophila embryos. When Drosophila tissue culture cells were prepared by a method which removes soluble protein and avoids fixation of the mitotic chromosomes, an enrichment for heterochromatin protein 1 in the heterochromatin of the chromosomes was discovered also.

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Pericentromeric Heterochromatin Domains Are Maintained without Accumulation of HP1

Molecular Biology of the Cell ◽

10.1091/mbc.e06-01-0025 ◽

2007 ◽

Vol 18 (4) ◽

pp. 1464-1471 ◽

Cited By ~ 23

Author(s):

Julio Mateos-Langerak ◽

Maartje C. Brink ◽

Martijn S. Luijsterburg ◽

Ineke van der Kraan ◽

Roel van Driel ◽

...

Keyword(s):

Histone H3 ◽

Dominant Negative ◽

Pericentromeric Heterochromatin ◽

Heterochromatin Protein 1 ◽

Dapi Staining ◽

Heterochromatin Protein ◽

3T3 Fibroblasts ◽

Heterochromatin Structure ◽

Hp1 Proteins

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 HP1α or HP1β proteins lacking a functional chromodomain. Expression of these truncated HP1 proteins individually or in combination resulted in a strong reduction of the accumulation of HP1α, HP1β, and HP1γ in pericentromeric heterochromatin domains in mouse 3T3 fibroblasts. The expression levels of HP1 did not change. The apparent displacement of HP1α, HP1β, and HP1γ 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 HP1α, HP1β, and HP1γ 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.

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Is HP1 an RNA detector that functions both in repression and activation?

The Journal of Cell Biology ◽

10.1083/jcb.200304145 ◽

2003 ◽

Vol 161 (4) ◽

pp. 671-672 ◽

Cited By ~ 9

Author(s):

Rebecca Kellum

Keyword(s):

Transcriptional Repression ◽

Polytene Chromosomes ◽

Pericentric Heterochromatin ◽

Original Study ◽

Heterochromatin Protein 1 ◽

Heterochromatin Protein ◽

Genetic Studies ◽

Transcript Levels ◽

Cytological Features ◽

Initial Discovery

Heterochromatin is defined as regions of compact chromatin that persist throughout the cell cycle (Heitz, 1928). The earliest cytological observations of heterochromatin were followed by ribonucleotide labeling experiments that showed it to be transcriptionally inert relative to the more typical euchromatic regions that decondense during interphase. Genetic studies of rearrangements that place euchromatic genes next to blocks of heterochromatin also pointed out the repressive nature of heterochromatin (Grigliatti, 1991; and references therein). The discovery of the heterochromatin-enriched protein heterochromatin protein 1 (HP1)**Abbreviation used in this paper: HP1, heterochromatin protein 1. by Elgin and co-workers in the mid-1980s suggested that the distinct cytological features of this chromatin may be related to its unique nucleoprotein composition (James and Elgin, 1986; James et al., 1989). HP1 immunostaining on polytene chromosomes from Drosophila larval salivary glands was used to show enrichment of the protein in pericentric heterochromatin. Since that initial discovery, HP1 homologues have been found in species ranging from fission yeast to humans where it is associated with gene silencing (Eissenberg and Elgin, 2000; and references therein). A number of euchromatic sites of localization were also reported in this original study. It has been generally assumed that these sites might constitute euchromatic sites of transcriptional repression by HP1. Indeed, several genes located at one of these sites (cytological region 31) have increased transcript levels in mutants for HP1 (Hwang et al., 2001).

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The Drosophila melanogaster dodecasatellite sequence is closely linked to the centromere and can form connections between sister chromatids during mitosis

Journal of Cell Science ◽

10.1242/jcs.105.1.41 ◽

1993 ◽

Vol 105 (1) ◽

pp. 41-50 ◽

Cited By ~ 4

Author(s):

M. Carmena ◽

J.P. Abad ◽

A. Villasante ◽

C. Gonzalez

Keyword(s):

Cell Cycle ◽

Drosophila Melanogaster ◽

In Situ Hybridisation ◽

Polytene Chromosomes ◽

Colchicine Treatment ◽

Dependent Manner ◽

Fluorescence In Situ Hybridisation ◽

Mitotic Chromosomes ◽

Sister Chromatids ◽

Diploid Cells

We have used fluorescence in situ hybridisation to wild-type and rearranged mitotic chromosomes to map the Drosophila melanogaster dodecasatellite sequence. It is located at a unique site, within the pericentric heterochromatin of the right arm of the third chromosome, closely linked to the primary constriction. In polytene chromosomes, dodecasatellite is found as one or a few dots in the central region of the chromocentre. In untreated diploid cells, dodecasatellite sequences are found as one or two dots throughout the cell cycle. This distribution can be altered in a cell cycle-dependent manner in two ways. Firstly, in interphase cells, hypotonic shock promotes the decondensation of the genomic region containing this satellite, resulting in a string-like structure. Secondly, some of the precociously separated sister chromatids produced by colchicine treatment show dodecasatellite within the intervening space connecting the main dodecasatellite signals of each chromatid. The distribution of dodecasatellite seems to be rather constant between individuals of the same species, as indicated by the lack of any detectable variations in its pattern amongst individuals from six geographically distant strains of D. melanogaster. On the other hand, the distribution of dodecasatellite shows a remarkable degree of variation amongst closely related species of the melanogaster subgroup ranging from a non-detectable signal in Drosophila yakuba and Drosophila teissieri, to staining in the X, second and third chromososomes of Drosophila mauritiana.

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Epigenetic displacement of HP1 from heterochromatin by HIV-1 Vpr causes premature sister chromatid separation

The Journal of Cell Biology ◽

10.1083/jcb.201010118 ◽

2011 ◽

Vol 194 (5) ◽

pp. 721-735 ◽

Cited By ~ 33

Author(s):

Mari Shimura ◽

Yusuke Toyoda ◽

Kenta Iijima ◽

Masanobu Kinomoto ◽

Kenzo Tokunaga ◽

...

Keyword(s):

Chromosome Segregation ◽

Mitotic Chromosomes ◽

Heterochromatin Protein 1 ◽

Heterochromatin Protein ◽

Viral Pathogen ◽

Chromatid Cohesion ◽

Epigenetic Disruption ◽

First Time ◽

Order Structures ◽

Hiv 1

Although pericentromeric heterochromatin is essential for chromosome segregation, its role in humans remains controversial. Dissecting the function of HIV-1–encoded Vpr, we unraveled important properties of heterochromatin during chromosome segregation. In Vpr-expressing cells, hRad21, hSgo1, and hMis12, which are crucial for proper chromosome segregation, were displaced from the centromeres of mitotic chromosomes, resulting in premature chromatid separation (PCS). Interestingly, Vpr displaced heterochromatin protein 1-α (HP1-α) and HP1-γ from chromatin. RNA interference (RNAi) experiments revealed that down-regulation of HP1-α and/or HP1-γ induced PCS, concomitant with the displacement of hRad21. Notably, Vpr stimulated the acetylation of histone H3, whereas p300 RNAi attenuated the Vpr-induced displacement of HP1-α and PCS. Furthermore, Vpr bound to p300 that was present in insoluble regions of the nucleus, suggesting that Vpr aberrantly recruits the histone acetyltransferase activity of p300 to chromatin, displaces HP1-α, and causes chromatid cohesion defects. Our study reveals for the first time centromere cohesion impairment resulting from epigenetic disruption of higher-order structures of heterochromatin by a viral pathogen.

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Heterochromatin protein 1 (HP1) is associated with induced gene expression in Drosophila euchromatin

The Journal of Cell Biology ◽

10.1083/jcb.200303012 ◽

2003 ◽

Vol 161 (4) ◽

pp. 707-714 ◽

Cited By ~ 141

Author(s):

Lucia Piacentini ◽

Laura Fanti ◽

Maria Berloco ◽

Barbara Perrini ◽

Sergio Pimpinelli

Keyword(s):

Heat Shock ◽

Polytene Chromosomes ◽

Gene Activity ◽

Chromosomal Protein ◽

Heterochromatin Protein 1 ◽

Heterochromatin Protein ◽

Heterochromatin Formation ◽

Telomere Capping ◽

Nonhistone Chromosomal Protein

Heterochromatin protein 1 (HP1) is a conserved nonhistone chromosomal protein, which is involved in heterochromatin formation and gene silencing in many organisms. In addition, it has been shown that HP1 is also involved in telomere capping in Drosophila. Here, we show a novel striking feature of this protein demonstrating its involvement in the activation of several euchromatic genes in Drosophila. By immunostaining experiments using an HP1 antibody, we found that HP1 is associated with developmental and heat shock–induced puffs on polytene chromosomes. Because the puffs are the cytological phenotype of intense gene activity, we did a detailed analysis of the heat shock–induced expression of the HSP70 encoding gene in larvae with different doses of HP1 and found that HP1 is positively involved in Hsp70 gene activity. These data significantly broaden the current views of the roles of HP1 in vivo by demonstrating that this protein has multifunctional roles.

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Loss of Heterochromatin Protein 1 Gamma Reduces the Number of Primordial Germ Cells via Impaired Cell Cycle Progression in Mice1

Biology of Reproduction ◽

10.1095/biolreprod.111.091512 ◽

2011 ◽

Vol 85 (5) ◽

pp. 1013-1024 ◽

Cited By ~ 28

Author(s):

Kanae Abe ◽

Chie Naruse ◽

Tomoaki Kato ◽

Takumi Nishiuchi ◽

Mitinori Saitou ◽

...

Keyword(s):

Cell Cycle ◽

Germ Cells ◽

Cell Cycle Progression ◽

Primordial Germ Cells ◽

Heterochromatin Protein 1 ◽

Cycle Progression ◽

Heterochromatin Protein

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Histone Hyperacetylation in Mitosis Prevents Sister Chromatid Separation and Produces Chromosome Segregation Defects

Molecular Biology of the Cell ◽

10.1091/mbc.e03-01-0860 ◽

2003 ◽

Vol 14 (9) ◽

pp. 3821-3833 ◽

Cited By ~ 106

Author(s):

Daniela Cimini ◽

Marta Mattiuzzo ◽

Liliana Torosantucci ◽

Francesca Degrassi

Keyword(s):

Sister Chromatid ◽

Histone Deacetylases ◽

Mitotic Checkpoint ◽

Chromosome Condensation ◽

Histone Deacetylation ◽

Centromeric Heterochromatin ◽

Chromatin Conformation ◽

Mitotic Chromosomes ◽

Heterochromatin Protein 1 ◽

Heterochromatin Protein

Posttranslational modifications of core histones contribute to driving changes in chromatin conformation and compaction. Herein, we investigated the role of histone deacetylation on the mitotic process by inhibiting histone deacetylases shortly before mitosis in human primary fibroblasts. Cells entering mitosis with hyperacetylated histones displayed altered chromatin conformation associated with decreased reactivity to the anti-Ser 10 phospho H3 antibody, increased recruitment of protein phosphatase 1-δ on mitotic chromosomes, and depletion of heterochromatin protein 1 from the centromeric heterochromatin. Inhibition of histone deacetylation before mitosis produced defective chromosome condensation and impaired mitotic progression in living cells, suggesting that improper chromosome condensation may induce mitotic checkpoint activation. In situ hybridization analysis on anaphase cells demonstrated the presence of chromatin bridges, which were caused by persisting cohesion along sister chromatid arms after centromere separation. Thus, the presence of hyperacetylated chromatin during mitosis impairs proper chromosome condensation during the pre-anaphase stages, resulting in poor sister chromatid resolution. Lagging chromosomes consisting of single or paired sisters were also induced by the presence of hyperacetylated histones, indicating that the less constrained centromeric organization associated with heterochromatin protein 1 depletion may promote the attachment of kinetochores to microtubules coming from both poles.

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Synthesis and Biological Evaluation of Novel Heterocyclic Imines Linked Coumarin- Thiazole Hybrids as Anticancer Agents

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520619666190207140120 ◽

2019 ◽

Vol 19 (4) ◽

pp. 557-566 ◽

Cited By ~ 7

Author(s):

Nerella S. Goud ◽

Mahammad S. Ghouse ◽

Jatoth Vishnu ◽

Jakkula Pranay ◽

Ravi Alvala ◽

...

Keyword(s):

Cell Cycle ◽

Membrane Potential ◽

Fluorescence Spectroscopy ◽

Mitochondrial Membrane Potential ◽

Mitochondrial Membrane ◽

Annexin V ◽

Biological Evaluation ◽

Dependent Manner ◽

Dapi Staining ◽

Dose Dependent

Background: Human Galectin-1, a protein of lectin family showing affinity towards β-galactosides has emerged as a critical regulator of tumor progression and metastasis, by modulating diverse biological events including homotypic cell aggregation, migration, apoptosis, angiogenesis and immune escape. Therefore, galectin-1 inhibitors might represent novel therapeutic agents for cancer. Methods: A new series of heterocyclic imines linked coumarin-thiazole hybrids (6a-6r) was synthesized and evaluated for its cytotoxic potential against a panel of six human cancer cell lines namely, lung (A549), prostate (DU-145), breast (MCF-7 & MDA-MB-231), colon (HCT-15 & HT-29) using MTT assay. Characteristic apoptotic assays like DAPI staining, cell cycle, annexin V and Mitochondrial membrane potential studies were performed for the most active compound. Furthermore, Gal-1 inhibition was confirmed by ELISA and fluorescence spectroscopy. Results: Among all, compound 6g 3-(2-(2-(pyridin-2-ylmethylene) hydrazineyl) thiazol-4-yl)-2H-chromen-2- one exhibited promising growth inhibition against HCT-15 colorectal cancer cells with an IC50 value of 1.28 ± 0.14 µM. The characteristic apoptotic morphological features like chromatin condensation, membrane blebbing and apoptotic body formation were clearly observed with compound 6g on HCT-15 cells using DAPI staining studies. Further, annexin V-FITC/PI assay confirmed effective early apoptosis induction by treatment with compound 6g. Loss of mitochondrial membrane potential and enhanced ROS generation were confirmed with JC-1 and DCFDA staining method, respectively by treatment with compound 6g, suggesting a possible mechanism for inducing apoptosis. Moreover, flow cytometric analysis revealed that compound 6g blocked G0/G1 phase of the cell cycle in a dose-dependent manner. Compound 6g effectively reduced the levels of Gal-1 protein in a dose-dependent manner. The binding constant (Ka) of 6g with Gal-1 was calculated from the intercept value which was observed as 1.9 x 107 M-1 by Fluorescence spectroscopy. Molecular docking studies showed strong interactions of compound 6g with Gal-1 protein. Conclusion: Our studies demonstrate the anticancer potential and Gal-1 inhibition of heterocyclic imines linked coumarin-thiazole hybrids.

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