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

Piacentini L, Fanti L, Berloco M, Perrini B, Pimpinelli S - J. Cell Biol. (2003)

Bottom Line: 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.These data significantly broaden the current views of the roles of HP1 in vivo by demonstrating that this protein has multifunctional roles.

View Article: PubMed Central - PubMed

Affiliation: Istituto Pasteur, Fondazione Cenci Bolognetti, Dipartimento di Genetica e Biologia Molecolare, Università di Roma La Sapienza, 00185 Roma, Italy.

ABSTRACT
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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Immunolocalization of HP1 on polytene chromosomes. The DAPI staining pattern and the HP1 immunopattern are shown in the top and middle panels, respectively. In the bottom panel, the merged patterns is shown. Intense signals are visible on the chromocenter (large arrowheads) and numerous signals are present along euchromatic arms and at all the telomeres (asterisks). Interestingly, the antibody also decorates several developmentally regulated puffs (arrows) including the ecdysone-induced puffs (small arrowheads). Note the absence of immunosignal in regions containing heat shock–inducing puffs (arrows in 87A and 87C).
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fig1: Immunolocalization of HP1 on polytene chromosomes. The DAPI staining pattern and the HP1 immunopattern are shown in the top and middle panels, respectively. In the bottom panel, the merged patterns is shown. Intense signals are visible on the chromocenter (large arrowheads) and numerous signals are present along euchromatic arms and at all the telomeres (asterisks). Interestingly, the antibody also decorates several developmentally regulated puffs (arrows) including the ecdysone-induced puffs (small arrowheads). Note the absence of immunosignal in regions containing heat shock–inducing puffs (arrows in 87A and 87C).

Mentions: Immunostaining of larval salivary gland chromosomes with an HP1 antibody revealed an enrichment of HP1 on the chromocenter, the fourth chromosome, and telomeres as already observed (James et al., 1989; Fanti et al., 1998; Fig. 1). In addition, the antibody also detected numerous sites along the euchromatic arms whose mapping is reported elsewhere (Fanti et al., 2003). Inspection of the specific loci to which HP1 binds revealed a striking result. Among the numerous euchromatic binding sites, we observed a localization of the protein to loci that form developmentally regulated chromosome puffs. We realized that, although not discussed, examples of puff staining with the HP1 antibody are evident also in a previous work by James et al. (1989). It is well-known that the puffs on polytene chromosomes of Drosophila and other Diptera are regions of high rates of RNA synthesis representing the visible expression of an intense gene activity at the chromosomal level. In salivary glands of third instar larvae, ∼10 prominent puffs are stably visible. During the late third instar larval and prepupal stages, the release of the hormone ecdysone into the hemolymph induces a sequence of puffing activity that involves ∼130 loci. Many of these loci have been mapped, and their characterization has shown that each puff has a specific temporal pattern of activity (Ashburner, 1972). As shown in Fig. 2 (A and B), three prominent ecdysone-induced puffs are clearly decorated by the HP1 antibody. This association is particularly suggestive of an involvement of HP1 in induced gene activity. To examine whether this result could be generalized, we asked if HP1 might also be found on other types of puffs such as those formed by transgenes under the control of heterologous promoters or heat shock–inducible puffs. We took advantage of the FLFW-1 transgenic Drosophila line characterized by Cavalli and Paro (1998)(1999). This strain contains the yeast transcriptional activator GAL-4 expressed under the control of the Hsp70 promoter and a GAL-4 activable UAS sequence that drives a lac-Z reporter gene. The reporter gene is flanked by Fab-7 and the mini-white gene. Previous characterization of the FLFW-1 insert by Cavalli and Paro (1998)(1999) showed that upon GAL-4 induction, the UAS-lac-Z gene forms a puff at the 61C9 region of the left arm of the third chromosome. As shown in Fig. 2 (C and D), we observed that, after induction of Gal-4, HP1 strongly accumulates on the puffed FLFW-1 insert.


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

Piacentini L, Fanti L, Berloco M, Perrini B, Pimpinelli S - J. Cell Biol. (2003)

Immunolocalization of HP1 on polytene chromosomes. The DAPI staining pattern and the HP1 immunopattern are shown in the top and middle panels, respectively. In the bottom panel, the merged patterns is shown. Intense signals are visible on the chromocenter (large arrowheads) and numerous signals are present along euchromatic arms and at all the telomeres (asterisks). Interestingly, the antibody also decorates several developmentally regulated puffs (arrows) including the ecdysone-induced puffs (small arrowheads). Note the absence of immunosignal in regions containing heat shock–inducing puffs (arrows in 87A and 87C).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2199350&req=5

fig1: Immunolocalization of HP1 on polytene chromosomes. The DAPI staining pattern and the HP1 immunopattern are shown in the top and middle panels, respectively. In the bottom panel, the merged patterns is shown. Intense signals are visible on the chromocenter (large arrowheads) and numerous signals are present along euchromatic arms and at all the telomeres (asterisks). Interestingly, the antibody also decorates several developmentally regulated puffs (arrows) including the ecdysone-induced puffs (small arrowheads). Note the absence of immunosignal in regions containing heat shock–inducing puffs (arrows in 87A and 87C).
Mentions: Immunostaining of larval salivary gland chromosomes with an HP1 antibody revealed an enrichment of HP1 on the chromocenter, the fourth chromosome, and telomeres as already observed (James et al., 1989; Fanti et al., 1998; Fig. 1). In addition, the antibody also detected numerous sites along the euchromatic arms whose mapping is reported elsewhere (Fanti et al., 2003). Inspection of the specific loci to which HP1 binds revealed a striking result. Among the numerous euchromatic binding sites, we observed a localization of the protein to loci that form developmentally regulated chromosome puffs. We realized that, although not discussed, examples of puff staining with the HP1 antibody are evident also in a previous work by James et al. (1989). It is well-known that the puffs on polytene chromosomes of Drosophila and other Diptera are regions of high rates of RNA synthesis representing the visible expression of an intense gene activity at the chromosomal level. In salivary glands of third instar larvae, ∼10 prominent puffs are stably visible. During the late third instar larval and prepupal stages, the release of the hormone ecdysone into the hemolymph induces a sequence of puffing activity that involves ∼130 loci. Many of these loci have been mapped, and their characterization has shown that each puff has a specific temporal pattern of activity (Ashburner, 1972). As shown in Fig. 2 (A and B), three prominent ecdysone-induced puffs are clearly decorated by the HP1 antibody. This association is particularly suggestive of an involvement of HP1 in induced gene activity. To examine whether this result could be generalized, we asked if HP1 might also be found on other types of puffs such as those formed by transgenes under the control of heterologous promoters or heat shock–inducible puffs. We took advantage of the FLFW-1 transgenic Drosophila line characterized by Cavalli and Paro (1998)(1999). This strain contains the yeast transcriptional activator GAL-4 expressed under the control of the Hsp70 promoter and a GAL-4 activable UAS sequence that drives a lac-Z reporter gene. The reporter gene is flanked by Fab-7 and the mini-white gene. Previous characterization of the FLFW-1 insert by Cavalli and Paro (1998)(1999) showed that upon GAL-4 induction, the UAS-lac-Z gene forms a puff at the 61C9 region of the left arm of the third chromosome. As shown in Fig. 2 (C and D), we observed that, after induction of Gal-4, HP1 strongly accumulates on the puffed FLFW-1 insert.

Bottom Line: 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.These data significantly broaden the current views of the roles of HP1 in vivo by demonstrating that this protein has multifunctional roles.

View Article: PubMed Central - PubMed

Affiliation: Istituto Pasteur, Fondazione Cenci Bolognetti, Dipartimento di Genetica e Biologia Molecolare, Università di Roma La Sapienza, 00185 Roma, Italy.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus