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Stress-induced transcription of satellite III repeats.

Jolly C, Metz A, Govin J, Vigneron M, Turner BM, Khochbin S, Vourc'h C - J. Cell Biol. (2003)

Bottom Line: These stress-induced structures, which form primarily on the 9q12 region in humans through direct binding of HSF1 to satellite III repeats, do not colocalize with transcription sites of known hsp genes.In this paper, we show that nuclear stress granules correspond to RNA polymerase II transcription factories where satellite III repeats are transcribed into large and stable RNAs that remain associated with the 9q12 region, even throughout mitosis.This work not only reveals the existence of a new major heat-induced transcript in human cells that may play a role in chromatin structure, but also provides evidence for a transcriptional activity within a locus considered so far as heterochromatic and silent.

View Article: PubMed Central - PubMed

Affiliation: INSERM U309, Institut A. Bonniot, 38706 La Tronche cedex, France. caroline.jolly@ujf-grenoble.fr

ABSTRACT
Exposure of mammalian cells to stress induces the activation of heat shock transcription factor 1 (HSF1) and the subsequent transcription of heat shock genes. Activation of the heat shock response also correlates with a rapid relocalization of HSF1 within a few nuclear structures termed nuclear stress granules. These stress-induced structures, which form primarily on the 9q12 region in humans through direct binding of HSF1 to satellite III repeats, do not colocalize with transcription sites of known hsp genes. In this paper, we show that nuclear stress granules correspond to RNA polymerase II transcription factories where satellite III repeats are transcribed into large and stable RNAs that remain associated with the 9q12 region, even throughout mitosis. This work not only reveals the existence of a new major heat-induced transcript in human cells that may play a role in chromatin structure, but also provides evidence for a transcriptional activity within a locus considered so far as heterochromatic and silent.

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RNA polymerase II is concentrated within nuclear stress granules. (A) RNA polymerase II (red) was detected by immunofluorescence together with HSF1 (green) in non–heat-shocked and heat-shocked HeLa cells. At 37°C, HSF1 is diffusely distributed in the nucleoplasm and cytoplasm while RNA polymerase II displays a fine nuclear punctate staining. After 1 h at 42°C, HSF1 is massively recruited to nuclear stress granules, and RNA polymerase II is also found accumulated in these structures (yellow in the merged image), in addition to a remaining diffuse staining of the nucleus. Bar, 5 μm. (B) Total cell extracts prepared from non–heat-shocked (37°C) or heat-shocked (1 h at 42°C) cells were analyzed by Western blot. The blot was sequentially detected with 7C2 anti-RNA polymerase II antibody (left) and with rabbit anti-HSF1 antibody (right). No cross-reactivity of the antibodies is observed. The lowered HSF1 mobility observed in the 42°C sample is due to stress-induced posttranslational modifications of the protein (for review see Pirkkala et al., 2001).
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fig1: RNA polymerase II is concentrated within nuclear stress granules. (A) RNA polymerase II (red) was detected by immunofluorescence together with HSF1 (green) in non–heat-shocked and heat-shocked HeLa cells. At 37°C, HSF1 is diffusely distributed in the nucleoplasm and cytoplasm while RNA polymerase II displays a fine nuclear punctate staining. After 1 h at 42°C, HSF1 is massively recruited to nuclear stress granules, and RNA polymerase II is also found accumulated in these structures (yellow in the merged image), in addition to a remaining diffuse staining of the nucleus. Bar, 5 μm. (B) Total cell extracts prepared from non–heat-shocked (37°C) or heat-shocked (1 h at 42°C) cells were analyzed by Western blot. The blot was sequentially detected with 7C2 anti-RNA polymerase II antibody (left) and with rabbit anti-HSF1 antibody (right). No cross-reactivity of the antibodies is observed. The lowered HSF1 mobility observed in the 42°C sample is due to stress-induced posttranslational modifications of the protein (for review see Pirkkala et al., 2001).

Mentions: We studied the possibility that nuclear stress granules could be sites of transcription by investigating by immunofluorescence the distribution in non–heat-shocked and heat-shocked cells of the key enzymes in transcription: RNA polymerases. Results are presented in Fig. 1. In contrast to RNA pol I and III (not depicted), the intranuclear distribution of RNA pol II detected with the 7C2 monoclonal antibody, which recognizes both nonphosphorylated and phosphorylated forms of the enzyme (Besse et al., 1995), was altered by heat shock. Indeed, after a 1-h heat shock, three to four nuclear foci were observed in most of the cells, in addition to a persisting diffuse staining of the nucleus and cytoplasm (Fig. 1 A). The colocalization of these pol II accumulation sites with nuclear stress granules was confirmed by codetection of HSF1 (Fig. 1 A). Similar observations were obtained with the MARA3 monoclonal antibody, which recognizes a phosphoepitope in the COOH-terminal domain of RNA pol II (Patturajan et al., 1998; not depicted). The specificity of both antibodies was assessed by Western blot on total cell extracts from non–heat-shocked and heat-shocked HeLa cells, and none of these antibodies was found to cross-react with HSF1 (Fig. 1 B).


Stress-induced transcription of satellite III repeats.

Jolly C, Metz A, Govin J, Vigneron M, Turner BM, Khochbin S, Vourc'h C - J. Cell Biol. (2003)

RNA polymerase II is concentrated within nuclear stress granules. (A) RNA polymerase II (red) was detected by immunofluorescence together with HSF1 (green) in non–heat-shocked and heat-shocked HeLa cells. At 37°C, HSF1 is diffusely distributed in the nucleoplasm and cytoplasm while RNA polymerase II displays a fine nuclear punctate staining. After 1 h at 42°C, HSF1 is massively recruited to nuclear stress granules, and RNA polymerase II is also found accumulated in these structures (yellow in the merged image), in addition to a remaining diffuse staining of the nucleus. Bar, 5 μm. (B) Total cell extracts prepared from non–heat-shocked (37°C) or heat-shocked (1 h at 42°C) cells were analyzed by Western blot. The blot was sequentially detected with 7C2 anti-RNA polymerase II antibody (left) and with rabbit anti-HSF1 antibody (right). No cross-reactivity of the antibodies is observed. The lowered HSF1 mobility observed in the 42°C sample is due to stress-induced posttranslational modifications of the protein (for review see Pirkkala et al., 2001).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171959&req=5

fig1: RNA polymerase II is concentrated within nuclear stress granules. (A) RNA polymerase II (red) was detected by immunofluorescence together with HSF1 (green) in non–heat-shocked and heat-shocked HeLa cells. At 37°C, HSF1 is diffusely distributed in the nucleoplasm and cytoplasm while RNA polymerase II displays a fine nuclear punctate staining. After 1 h at 42°C, HSF1 is massively recruited to nuclear stress granules, and RNA polymerase II is also found accumulated in these structures (yellow in the merged image), in addition to a remaining diffuse staining of the nucleus. Bar, 5 μm. (B) Total cell extracts prepared from non–heat-shocked (37°C) or heat-shocked (1 h at 42°C) cells were analyzed by Western blot. The blot was sequentially detected with 7C2 anti-RNA polymerase II antibody (left) and with rabbit anti-HSF1 antibody (right). No cross-reactivity of the antibodies is observed. The lowered HSF1 mobility observed in the 42°C sample is due to stress-induced posttranslational modifications of the protein (for review see Pirkkala et al., 2001).
Mentions: We studied the possibility that nuclear stress granules could be sites of transcription by investigating by immunofluorescence the distribution in non–heat-shocked and heat-shocked cells of the key enzymes in transcription: RNA polymerases. Results are presented in Fig. 1. In contrast to RNA pol I and III (not depicted), the intranuclear distribution of RNA pol II detected with the 7C2 monoclonal antibody, which recognizes both nonphosphorylated and phosphorylated forms of the enzyme (Besse et al., 1995), was altered by heat shock. Indeed, after a 1-h heat shock, three to four nuclear foci were observed in most of the cells, in addition to a persisting diffuse staining of the nucleus and cytoplasm (Fig. 1 A). The colocalization of these pol II accumulation sites with nuclear stress granules was confirmed by codetection of HSF1 (Fig. 1 A). Similar observations were obtained with the MARA3 monoclonal antibody, which recognizes a phosphoepitope in the COOH-terminal domain of RNA pol II (Patturajan et al., 1998; not depicted). The specificity of both antibodies was assessed by Western blot on total cell extracts from non–heat-shocked and heat-shocked HeLa cells, and none of these antibodies was found to cross-react with HSF1 (Fig. 1 B).

Bottom Line: These stress-induced structures, which form primarily on the 9q12 region in humans through direct binding of HSF1 to satellite III repeats, do not colocalize with transcription sites of known hsp genes.In this paper, we show that nuclear stress granules correspond to RNA polymerase II transcription factories where satellite III repeats are transcribed into large and stable RNAs that remain associated with the 9q12 region, even throughout mitosis.This work not only reveals the existence of a new major heat-induced transcript in human cells that may play a role in chromatin structure, but also provides evidence for a transcriptional activity within a locus considered so far as heterochromatic and silent.

View Article: PubMed Central - PubMed

Affiliation: INSERM U309, Institut A. Bonniot, 38706 La Tronche cedex, France. caroline.jolly@ujf-grenoble.fr

ABSTRACT
Exposure of mammalian cells to stress induces the activation of heat shock transcription factor 1 (HSF1) and the subsequent transcription of heat shock genes. Activation of the heat shock response also correlates with a rapid relocalization of HSF1 within a few nuclear structures termed nuclear stress granules. These stress-induced structures, which form primarily on the 9q12 region in humans through direct binding of HSF1 to satellite III repeats, do not colocalize with transcription sites of known hsp genes. In this paper, we show that nuclear stress granules correspond to RNA polymerase II transcription factories where satellite III repeats are transcribed into large and stable RNAs that remain associated with the 9q12 region, even throughout mitosis. This work not only reveals the existence of a new major heat-induced transcript in human cells that may play a role in chromatin structure, but also provides evidence for a transcriptional activity within a locus considered so far as heterochromatic and silent.

Show MeSH
Related in: MedlinePlus