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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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Stress-induced sat III transcription is HSF1 dependent. The GFP-tagged DBF+TRIM mutant of HSF1 (green) was transiently transfected into HeLa cells and codetected with either (A) endogenous HSF1, (B) transiently expressed CBP-HA, (C) acetylated histone H4, (D) RNA polymerase II, or (E) pHuR98 transcripts (red). Overexpression of this mutant prevents the accumulation of endogenous HSF1, acetylated H4, CBP-HA, and RNA polymerase II into the granules, both at 37°C and at 42°C. Likewise, pHuR98 transcripts are not detected in the granules formed by the HSF1 mutant at 37°C and 42°C. Arrowheads in D point to granules of RNA pol II. (F) HeLa cells were transiently transfected with a plasmid coding for the HSP70 chaperone. HSP70 was then detected by immunofluorescence together with either the endogenous HSF1, transiently coexpressed CBP-HA, or acetylated histone H4 detected by immunofluorescence, or with pHuR98 transcripts revealed by RNA FISH. Only images of cells exposed for 1 h at 42°C are shown. In HSP70-overexpressing cells, the endogenous HSF1, CBP-HA, and acetylated histone H4 are not recruited to nuclear stress granules, and pHuR98 transcript foci are not present. The white lines in E and F indicate that the cells were taken from different fields. Bars, 10 μm.
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fig6: Stress-induced sat III transcription is HSF1 dependent. The GFP-tagged DBF+TRIM mutant of HSF1 (green) was transiently transfected into HeLa cells and codetected with either (A) endogenous HSF1, (B) transiently expressed CBP-HA, (C) acetylated histone H4, (D) RNA polymerase II, or (E) pHuR98 transcripts (red). Overexpression of this mutant prevents the accumulation of endogenous HSF1, acetylated H4, CBP-HA, and RNA polymerase II into the granules, both at 37°C and at 42°C. Likewise, pHuR98 transcripts are not detected in the granules formed by the HSF1 mutant at 37°C and 42°C. Arrowheads in D point to granules of RNA pol II. (F) HeLa cells were transiently transfected with a plasmid coding for the HSP70 chaperone. HSP70 was then detected by immunofluorescence together with either the endogenous HSF1, transiently coexpressed CBP-HA, or acetylated histone H4 detected by immunofluorescence, or with pHuR98 transcripts revealed by RNA FISH. Only images of cells exposed for 1 h at 42°C are shown. In HSP70-overexpressing cells, the endogenous HSF1, CBP-HA, and acetylated histone H4 are not recruited to nuclear stress granules, and pHuR98 transcript foci are not present. The white lines in E and F indicate that the cells were taken from different fields. Bars, 10 μm.

Mentions: The question of whether HSF1 is the key determinant in the induced transcription of sat III repeats was first addressed by using the DBD+TRIM deletion mutant of HSF1, which only contains the DNA-binding and trimerization domains and forms granules constitutively (Jolly et al., 2002). Interestingly, this mutant acts as a dominant negative, by preventing the endogenous HSF1 from accumulating into the granules upon heat shock (Fig. 6 A). We found that acetylated histone H4, CBP-HA, RNA polymerase II, and sat III transcripts were never present in the granules formed by the DBD+TRIM mutant, both at 37°C and at 42°C (Fig. 6, B–E). As a confirmation to this, we also prevented the targeting of HSF1 to the granules by overexpressing HSP70, which is a negative regulator of HSF1 activity (Shi et al., 1998). As expected, upon heat shock, HSP70 overexpression prevented the accumulation of HSF1, acetylated histones, and CBP-HA in the granules, and the subsequent transcription of sat III repeats (Fig. 6 F).


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)

Stress-induced sat III transcription is HSF1 dependent. The GFP-tagged DBF+TRIM mutant of HSF1 (green) was transiently transfected into HeLa cells and codetected with either (A) endogenous HSF1, (B) transiently expressed CBP-HA, (C) acetylated histone H4, (D) RNA polymerase II, or (E) pHuR98 transcripts (red). Overexpression of this mutant prevents the accumulation of endogenous HSF1, acetylated H4, CBP-HA, and RNA polymerase II into the granules, both at 37°C and at 42°C. Likewise, pHuR98 transcripts are not detected in the granules formed by the HSF1 mutant at 37°C and 42°C. Arrowheads in D point to granules of RNA pol II. (F) HeLa cells were transiently transfected with a plasmid coding for the HSP70 chaperone. HSP70 was then detected by immunofluorescence together with either the endogenous HSF1, transiently coexpressed CBP-HA, or acetylated histone H4 detected by immunofluorescence, or with pHuR98 transcripts revealed by RNA FISH. Only images of cells exposed for 1 h at 42°C are shown. In HSP70-overexpressing cells, the endogenous HSF1, CBP-HA, and acetylated histone H4 are not recruited to nuclear stress granules, and pHuR98 transcript foci are not present. The white lines in E and F indicate that the cells were taken from different fields. Bars, 10 μm.
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Related In: Results  -  Collection

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fig6: Stress-induced sat III transcription is HSF1 dependent. The GFP-tagged DBF+TRIM mutant of HSF1 (green) was transiently transfected into HeLa cells and codetected with either (A) endogenous HSF1, (B) transiently expressed CBP-HA, (C) acetylated histone H4, (D) RNA polymerase II, or (E) pHuR98 transcripts (red). Overexpression of this mutant prevents the accumulation of endogenous HSF1, acetylated H4, CBP-HA, and RNA polymerase II into the granules, both at 37°C and at 42°C. Likewise, pHuR98 transcripts are not detected in the granules formed by the HSF1 mutant at 37°C and 42°C. Arrowheads in D point to granules of RNA pol II. (F) HeLa cells were transiently transfected with a plasmid coding for the HSP70 chaperone. HSP70 was then detected by immunofluorescence together with either the endogenous HSF1, transiently coexpressed CBP-HA, or acetylated histone H4 detected by immunofluorescence, or with pHuR98 transcripts revealed by RNA FISH. Only images of cells exposed for 1 h at 42°C are shown. In HSP70-overexpressing cells, the endogenous HSF1, CBP-HA, and acetylated histone H4 are not recruited to nuclear stress granules, and pHuR98 transcript foci are not present. The white lines in E and F indicate that the cells were taken from different fields. Bars, 10 μm.
Mentions: The question of whether HSF1 is the key determinant in the induced transcription of sat III repeats was first addressed by using the DBD+TRIM deletion mutant of HSF1, which only contains the DNA-binding and trimerization domains and forms granules constitutively (Jolly et al., 2002). Interestingly, this mutant acts as a dominant negative, by preventing the endogenous HSF1 from accumulating into the granules upon heat shock (Fig. 6 A). We found that acetylated histone H4, CBP-HA, RNA polymerase II, and sat III transcripts were never present in the granules formed by the DBD+TRIM mutant, both at 37°C and at 42°C (Fig. 6, B–E). As a confirmation to this, we also prevented the targeting of HSF1 to the granules by overexpressing HSP70, which is a negative regulator of HSF1 activity (Shi et al., 1998). As expected, upon heat shock, HSP70 overexpression prevented the accumulation of HSF1, acetylated histones, and CBP-HA in the granules, and the subsequent transcription of sat III repeats (Fig. 6 F).

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