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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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The RNase treatment of wild-type polytene chromosomes and the Su(var)2-502 HP1 mutation affects the euchromatic HP1 binding with similar immunopatterns observed. (A) HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of untreated wild-type larvae. Many of euchromatic HP1 immunosignals apparent in the controls are very faint or not visible at all in the RNase-treated case. Interestingly, the physiological puffs also lack HP1 immunosignals. The few immunosignals still visible, such as the signal on the 60B region (arrow) of the second chromosome and on the 14A and 14C regions of the X chromosome (arrows), are the same present in the polytene chromosomes of wild-type larvae after the heat shock treatment reported in Fig. 2 (F and G). Note that the immunostaining on the telomeres (asterisks), the chromocenter, the fourth chromosome, and the 31 region does not appear to be affected (large arrowheads). (B) A higher magnification showing HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of heat-shocked wild-type larvae. The heat shock–induced puffs in 87A and 87C do not show immunostaining (arrows). The arrowhead points to one of the few residual immunosignals. (C) HP1 immunopattern on polytene chromosomes from Su(var)2-502/Su(var)2-505 mutant larvae. The chromocenter, the telomeres, and the 31 region (arrowheads) do not seem to be strongly affected, whereas the euchromatic immunopattern appears identical to that observed after RNase treatment showing only the same few immunosignals on 60B, 14A, and 14C (arrows). (D) A higher magnification view of an HP1-immunostained polytene chromosomes from salivary glands of heat-shocked Su(var)2-502/Su(var)2-505 mutant larvae. The heat shock–induced puffs at 87A and 87C are not immunostained (arrows). Note that in this case the same residual immunosignal, visible in B, is also present (arrowhead).
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fig5: The RNase treatment of wild-type polytene chromosomes and the Su(var)2-502 HP1 mutation affects the euchromatic HP1 binding with similar immunopatterns observed. (A) HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of untreated wild-type larvae. Many of euchromatic HP1 immunosignals apparent in the controls are very faint or not visible at all in the RNase-treated case. Interestingly, the physiological puffs also lack HP1 immunosignals. The few immunosignals still visible, such as the signal on the 60B region (arrow) of the second chromosome and on the 14A and 14C regions of the X chromosome (arrows), are the same present in the polytene chromosomes of wild-type larvae after the heat shock treatment reported in Fig. 2 (F and G). Note that the immunostaining on the telomeres (asterisks), the chromocenter, the fourth chromosome, and the 31 region does not appear to be affected (large arrowheads). (B) A higher magnification showing HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of heat-shocked wild-type larvae. The heat shock–induced puffs in 87A and 87C do not show immunostaining (arrows). The arrowhead points to one of the few residual immunosignals. (C) HP1 immunopattern on polytene chromosomes from Su(var)2-502/Su(var)2-505 mutant larvae. The chromocenter, the telomeres, and the 31 region (arrowheads) do not seem to be strongly affected, whereas the euchromatic immunopattern appears identical to that observed after RNase treatment showing only the same few immunosignals on 60B, 14A, and 14C (arrows). (D) A higher magnification view of an HP1-immunostained polytene chromosomes from salivary glands of heat-shocked Su(var)2-502/Su(var)2-505 mutant larvae. The heat shock–induced puffs at 87A and 87C are not immunostained (arrows). Note that in this case the same residual immunosignal, visible in B, is also present (arrowhead).

Mentions: The chromatin IP studies indicated an enrichment of HP1 in the coding region of Hsp70, but did not indicate whether HP1 binding is dependent on the presence of RNA. To test this point, we treated polytene chromosomes of untreated and heat-shocked wild-type larvae with RNase, and fixed the chromosomes followed by immunostaining with anti-HP1 antibody. Fig. 5 A shows the results of the RNase treatment on HP1 staining in nonheat-shocked polytenes. We observed differences in the effects of the RNase treatment depending on the chromosomal region. A loss of HP1 immunosignals was evident at many euchromatic sites with a pattern very similar, if not identical, to the immunopattern observed after heat shock in RNase untreated chromosomes (Fig. 2 F). However, the RNase treatment did not affect the immunofluorescence on the chromocenter, telomeres, and the 31 region. In addition, we observed that RNase treatment results in the removal of HP1 at the heat shock–induced puffs (Fig. 5 B). We also found that HP1 is not recruited to the heat shock puffs when they are induced by sodium salicylate (unpublished data). This substance is known to induce heat shock puff formation without RNA transcription (Winegarden et al., 1996) and, therefore, our observation strongly suggests that HP1 recruitment to the puffs depends on the presence of RNA transcripts.


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)

The RNase treatment of wild-type polytene chromosomes and the Su(var)2-502 HP1 mutation affects the euchromatic HP1 binding with similar immunopatterns observed. (A) HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of untreated wild-type larvae. Many of euchromatic HP1 immunosignals apparent in the controls are very faint or not visible at all in the RNase-treated case. Interestingly, the physiological puffs also lack HP1 immunosignals. The few immunosignals still visible, such as the signal on the 60B region (arrow) of the second chromosome and on the 14A and 14C regions of the X chromosome (arrows), are the same present in the polytene chromosomes of wild-type larvae after the heat shock treatment reported in Fig. 2 (F and G). Note that the immunostaining on the telomeres (asterisks), the chromocenter, the fourth chromosome, and the 31 region does not appear to be affected (large arrowheads). (B) A higher magnification showing HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of heat-shocked wild-type larvae. The heat shock–induced puffs in 87A and 87C do not show immunostaining (arrows). The arrowhead points to one of the few residual immunosignals. (C) HP1 immunopattern on polytene chromosomes from Su(var)2-502/Su(var)2-505 mutant larvae. The chromocenter, the telomeres, and the 31 region (arrowheads) do not seem to be strongly affected, whereas the euchromatic immunopattern appears identical to that observed after RNase treatment showing only the same few immunosignals on 60B, 14A, and 14C (arrows). (D) A higher magnification view of an HP1-immunostained polytene chromosomes from salivary glands of heat-shocked Su(var)2-502/Su(var)2-505 mutant larvae. The heat shock–induced puffs at 87A and 87C are not immunostained (arrows). Note that in this case the same residual immunosignal, visible in B, is also present (arrowhead).
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fig5: The RNase treatment of wild-type polytene chromosomes and the Su(var)2-502 HP1 mutation affects the euchromatic HP1 binding with similar immunopatterns observed. (A) HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of untreated wild-type larvae. Many of euchromatic HP1 immunosignals apparent in the controls are very faint or not visible at all in the RNase-treated case. Interestingly, the physiological puffs also lack HP1 immunosignals. The few immunosignals still visible, such as the signal on the 60B region (arrow) of the second chromosome and on the 14A and 14C regions of the X chromosome (arrows), are the same present in the polytene chromosomes of wild-type larvae after the heat shock treatment reported in Fig. 2 (F and G). Note that the immunostaining on the telomeres (asterisks), the chromocenter, the fourth chromosome, and the 31 region does not appear to be affected (large arrowheads). (B) A higher magnification showing HP1 immunostaining of RNase-treated polytene chromosomes from salivary glands of heat-shocked wild-type larvae. The heat shock–induced puffs in 87A and 87C do not show immunostaining (arrows). The arrowhead points to one of the few residual immunosignals. (C) HP1 immunopattern on polytene chromosomes from Su(var)2-502/Su(var)2-505 mutant larvae. The chromocenter, the telomeres, and the 31 region (arrowheads) do not seem to be strongly affected, whereas the euchromatic immunopattern appears identical to that observed after RNase treatment showing only the same few immunosignals on 60B, 14A, and 14C (arrows). (D) A higher magnification view of an HP1-immunostained polytene chromosomes from salivary glands of heat-shocked Su(var)2-502/Su(var)2-505 mutant larvae. The heat shock–induced puffs at 87A and 87C are not immunostained (arrows). Note that in this case the same residual immunosignal, visible in B, is also present (arrowhead).
Mentions: The chromatin IP studies indicated an enrichment of HP1 in the coding region of Hsp70, but did not indicate whether HP1 binding is dependent on the presence of RNA. To test this point, we treated polytene chromosomes of untreated and heat-shocked wild-type larvae with RNase, and fixed the chromosomes followed by immunostaining with anti-HP1 antibody. Fig. 5 A shows the results of the RNase treatment on HP1 staining in nonheat-shocked polytenes. We observed differences in the effects of the RNase treatment depending on the chromosomal region. A loss of HP1 immunosignals was evident at many euchromatic sites with a pattern very similar, if not identical, to the immunopattern observed after heat shock in RNase untreated chromosomes (Fig. 2 F). However, the RNase treatment did not affect the immunofluorescence on the chromocenter, telomeres, and the 31 region. In addition, we observed that RNase treatment results in the removal of HP1 at the heat shock–induced puffs (Fig. 5 B). We also found that HP1 is not recruited to the heat shock puffs when they are induced by sodium salicylate (unpublished data). This substance is known to induce heat shock puff formation without RNA transcription (Winegarden et al., 1996) and, therefore, our observation strongly suggests that HP1 recruitment to the puffs depends on the presence of RNA transcripts.

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