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HDAC6-ubiquitin interaction controls the duration of HSF1 activation after heat shock.

Pernet L, Faure V, Gilquin B, Dufour-Guérin S, Khochbin S, Vourc'h C - Mol. Biol. Cell (2014)

Bottom Line: Here we show that a full response to heat shock (activation of both HSP70 and HSP25) depends on the duration of HSF1 activation, which is itself controlled by HDAC6, a unique deacetylase known to bind monoubiquitin and polyubiquitin with high affinity.In cells expressing HDAC6 mutated in the ubiquitin-binding domain, the AAA ATPase factor p97/VCP mediates rapid inactivation of HSF1, precluding late activation of the HSP25 gene.In these cells, knockdown of p97/VCP rescues HSF1 from this rapid inactivation and restores HSP25 expression.

View Article: PubMed Central - PubMed

Affiliation: University Grenoble-Alpes, CRI INSERM, U823, Institut Albert Bonniot, La Tronche 38042, Grenoble Cedex 9, France.

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HDAC6 mutated in its ZnF-UBP domain increases the percentage of heat-induced apoptosis and prevents HSP25 accumulation in heat-shocked cells. (A) WT and HDAC6 mutant– expressing cells were submitted or not (NHS) to heat shock, followed by different times of recovery (Rec). The percentage of apoptotic cells was quantified by fluorescence-activated cell sorting through caspase 3 detection. Histograms represent average values determined from two independent experiments. The results were normalized with the wild-type point in unstressed conditions. (B) 3T3 cells were transiently transfected with the indicated HA-tagged HDAC6 expression vectors and submitted to a recovery period of 24 h after heat shock. Transfected cells were detected using an anti-HA antibody. (C) The proportion of cells expressing endogenous HSP25 was calculated using an anti-HSP25 antibody. The histograms represent values from three independent experiments.
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Figure 6: HDAC6 mutated in its ZnF-UBP domain increases the percentage of heat-induced apoptosis and prevents HSP25 accumulation in heat-shocked cells. (A) WT and HDAC6 mutant– expressing cells were submitted or not (NHS) to heat shock, followed by different times of recovery (Rec). The percentage of apoptotic cells was quantified by fluorescence-activated cell sorting through caspase 3 detection. Histograms represent average values determined from two independent experiments. The results were normalized with the wild-type point in unstressed conditions. (B) 3T3 cells were transiently transfected with the indicated HA-tagged HDAC6 expression vectors and submitted to a recovery period of 24 h after heat shock. Transfected cells were detected using an anti-HA antibody. (C) The proportion of cells expressing endogenous HSP25 was calculated using an anti-HSP25 antibody. The histograms represent values from three independent experiments.

Mentions: The ubiquitin-binding domain of HDAC6 protects cells against stress-induced apoptosis. Because HSP25 is well known for its role in cell protection against stress-induced apoptosis (Huot et al., 1991; Lee et al., 2004), we next sought to determine whether down-regulation of HSP25 in Ubm HDAC6–expressing cells would have a negative effect on cell protection against heat-induced apoptosis. We thus measured and compared the percentage of apoptotic cells in wild-type cells and in HDAC6 mutant–expressing cells submitted or not to a heat shock followed by recovery of 12 and 24 h at 37°C (Figure 6A). As expected, the percentage of apoptotic cells 24 h after heat shock was significantly higher (35-fold increase) in HDAC6 Ubm–expressing cells than in unstressed wild-type cells and three times higher than in all other cell lines at the same time point. These results confirm the importance of the ubiquitin-binding domain of HDAC6 in the HSP25-mediated protection of cells against cell death.


HDAC6-ubiquitin interaction controls the duration of HSF1 activation after heat shock.

Pernet L, Faure V, Gilquin B, Dufour-Guérin S, Khochbin S, Vourc'h C - Mol. Biol. Cell (2014)

HDAC6 mutated in its ZnF-UBP domain increases the percentage of heat-induced apoptosis and prevents HSP25 accumulation in heat-shocked cells. (A) WT and HDAC6 mutant– expressing cells were submitted or not (NHS) to heat shock, followed by different times of recovery (Rec). The percentage of apoptotic cells was quantified by fluorescence-activated cell sorting through caspase 3 detection. Histograms represent average values determined from two independent experiments. The results were normalized with the wild-type point in unstressed conditions. (B) 3T3 cells were transiently transfected with the indicated HA-tagged HDAC6 expression vectors and submitted to a recovery period of 24 h after heat shock. Transfected cells were detected using an anti-HA antibody. (C) The proportion of cells expressing endogenous HSP25 was calculated using an anti-HSP25 antibody. The histograms represent values from three independent experiments.
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Figure 6: HDAC6 mutated in its ZnF-UBP domain increases the percentage of heat-induced apoptosis and prevents HSP25 accumulation in heat-shocked cells. (A) WT and HDAC6 mutant– expressing cells were submitted or not (NHS) to heat shock, followed by different times of recovery (Rec). The percentage of apoptotic cells was quantified by fluorescence-activated cell sorting through caspase 3 detection. Histograms represent average values determined from two independent experiments. The results were normalized with the wild-type point in unstressed conditions. (B) 3T3 cells were transiently transfected with the indicated HA-tagged HDAC6 expression vectors and submitted to a recovery period of 24 h after heat shock. Transfected cells were detected using an anti-HA antibody. (C) The proportion of cells expressing endogenous HSP25 was calculated using an anti-HSP25 antibody. The histograms represent values from three independent experiments.
Mentions: The ubiquitin-binding domain of HDAC6 protects cells against stress-induced apoptosis. Because HSP25 is well known for its role in cell protection against stress-induced apoptosis (Huot et al., 1991; Lee et al., 2004), we next sought to determine whether down-regulation of HSP25 in Ubm HDAC6–expressing cells would have a negative effect on cell protection against heat-induced apoptosis. We thus measured and compared the percentage of apoptotic cells in wild-type cells and in HDAC6 mutant–expressing cells submitted or not to a heat shock followed by recovery of 12 and 24 h at 37°C (Figure 6A). As expected, the percentage of apoptotic cells 24 h after heat shock was significantly higher (35-fold increase) in HDAC6 Ubm–expressing cells than in unstressed wild-type cells and three times higher than in all other cell lines at the same time point. These results confirm the importance of the ubiquitin-binding domain of HDAC6 in the HSP25-mediated protection of cells against cell death.

Bottom Line: Here we show that a full response to heat shock (activation of both HSP70 and HSP25) depends on the duration of HSF1 activation, which is itself controlled by HDAC6, a unique deacetylase known to bind monoubiquitin and polyubiquitin with high affinity.In cells expressing HDAC6 mutated in the ubiquitin-binding domain, the AAA ATPase factor p97/VCP mediates rapid inactivation of HSF1, precluding late activation of the HSP25 gene.In these cells, knockdown of p97/VCP rescues HSF1 from this rapid inactivation and restores HSP25 expression.

View Article: PubMed Central - PubMed

Affiliation: University Grenoble-Alpes, CRI INSERM, U823, Institut Albert Bonniot, La Tronche 38042, Grenoble Cedex 9, France.

Show MeSH
Related in: MedlinePlus