Limits...
Programmed cell death 5 mediates HDAC3 decay to promote genotoxic stress response.

Choi HK, Choi Y, Park ES, Park SY, Lee SH, Seo J, Jeong MH, Jeong JW, Jeong JH, Lee PC, Choi KC, Yoon HG - Nat Commun (2015)

Bottom Line: Restoration of PDCD5(WT) in PDCD5(-/-) MEFs restores ET-induced HDAC3 cleavage.Reduction of both PDCD5 and p53, but not reduction of either protein alone, significantly enhances in vivo tumorigenicity of AGS gastric cancer cells and correlates with poor prognosis in gastric cancer patients.Our results define a mechanism for p53 activation via PDCD5-dependent HDAC3 decay under genotoxic stress conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul 120-752, Korea.

ABSTRACT
The inhibition of p53 activity by histone deacetylase 3 (HDAC3) has been reported, but the precise molecular mechanism is unknown. Here we show that programmed cell death 5 (PDCD5) selectively mediates HDAC3 dissociation from p53, which induces HDAC3 cleavage and ubiquitin-dependent proteasomal degradation. Casein kinase 2 alpha phosphorylates PDCD5 at Ser-119 to enhance its stability and importin 13-mediated nuclear translocation of PDCD5. Genetic deletion of PDCD5 abrogates etoposide (ET)-induced p53 stabilization and HDAC3 cleavage, indicating an essential role of PDCD5 in p53 activation. Restoration of PDCD5(WT) in PDCD5(-/-) MEFs restores ET-induced HDAC3 cleavage. Reduction of both PDCD5 and p53, but not reduction of either protein alone, significantly enhances in vivo tumorigenicity of AGS gastric cancer cells and correlates with poor prognosis in gastric cancer patients. Our results define a mechanism for p53 activation via PDCD5-dependent HDAC3 decay under genotoxic stress conditions.

No MeSH data available.


Related in: MedlinePlus

PDCD5 stabilizes p53 by inducing dissociation of the HDAC3–p53 complex and concomitantly triggering cytosolic cleavage of HDAC3.(a) Overexpression of PDCD5 reduces the activity of HDAC3. HCT-116 cells were transfected with the indicated plasmids. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and then HDAC3 activity was measured. Error bars, s.d. (n=3). *P<0.05. (b) Depletion of PDCD5 abolishes the ET-induced reduction of HDAC3 activity. Cells were transfected with the indicated siRNAs. Cells were treated with ET (100 μM, 12 h) or STS (1 μM, 12 h) and assayed for HDAC3 activity. Error bars, s.d. (n=3). *P<0.05. (c) Knockdown of PDCD5 abrogates the association of HDAC3 with active caspase-3 in response to ET treatment. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and subsequently immunoblotted with the indicated antibodies. Arrow indicates cleaved HDAC3. (d) Knockdown of PDCD5 prevents cytoplasmic cleavage of HDAC3 in response to ET treatment. Following cell fractionation, fractions were immunoblotted with the indicated antibodies. (e) Overexpression of PDCD5 enhances ET-induced cytoplasmic translocation of HDAC3. Immunofluorescence analysis was performed as described in the Supplementary Experimental Procedures Section. Representative images of three independent experiments are shown. (f) PDCD5 knockdown diminishes the ET-induced dissociation of HDAC3 from p53. Indicated shRNA-expressing HCT-116 cells were treated with ET. Whole-cell lysates were analysed by western blotting with the indicated antibodies. (g) Overexpression of PDCD5 dissociates HDAC3 from p53. Cells were transfected with PDCD5 plasmids. Immunoprecipitation and immunoblotting analyses were performed with the indicated antibodies. (h,i) PDCD5 increases p53 acetylation and stability via mediating HDAC3 cleavage. Cells were treated with ET (h) or cycloheximide (i) and subsequently immunoblotted with the indicated antibodies. Scale bar, 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4490383&req=5

f2: PDCD5 stabilizes p53 by inducing dissociation of the HDAC3–p53 complex and concomitantly triggering cytosolic cleavage of HDAC3.(a) Overexpression of PDCD5 reduces the activity of HDAC3. HCT-116 cells were transfected with the indicated plasmids. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and then HDAC3 activity was measured. Error bars, s.d. (n=3). *P<0.05. (b) Depletion of PDCD5 abolishes the ET-induced reduction of HDAC3 activity. Cells were transfected with the indicated siRNAs. Cells were treated with ET (100 μM, 12 h) or STS (1 μM, 12 h) and assayed for HDAC3 activity. Error bars, s.d. (n=3). *P<0.05. (c) Knockdown of PDCD5 abrogates the association of HDAC3 with active caspase-3 in response to ET treatment. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and subsequently immunoblotted with the indicated antibodies. Arrow indicates cleaved HDAC3. (d) Knockdown of PDCD5 prevents cytoplasmic cleavage of HDAC3 in response to ET treatment. Following cell fractionation, fractions were immunoblotted with the indicated antibodies. (e) Overexpression of PDCD5 enhances ET-induced cytoplasmic translocation of HDAC3. Immunofluorescence analysis was performed as described in the Supplementary Experimental Procedures Section. Representative images of three independent experiments are shown. (f) PDCD5 knockdown diminishes the ET-induced dissociation of HDAC3 from p53. Indicated shRNA-expressing HCT-116 cells were treated with ET. Whole-cell lysates were analysed by western blotting with the indicated antibodies. (g) Overexpression of PDCD5 dissociates HDAC3 from p53. Cells were transfected with PDCD5 plasmids. Immunoprecipitation and immunoblotting analyses were performed with the indicated antibodies. (h,i) PDCD5 increases p53 acetylation and stability via mediating HDAC3 cleavage. Cells were treated with ET (h) or cycloheximide (i) and subsequently immunoblotted with the indicated antibodies. Scale bar, 10 μm.

Mentions: Given results suggesting PDCD5 is involved in caspase-3-dependent HDAC3 cleavage, we next investigated the role of PDCD5 in regulation of HDAC3 function. Because HDAC activity is known to be a critical function of HDAC3 (ref. 26), we first assessed the change in HDAC3 activity after either overexpression or depletion of PDCD5 in cells. An increase in wild-type PDCD5 expression, but not mutant PDCD5L6R, resulted in a decrease of HDAC3 histone deacetylase activity (Fig. 2a). However, overexpression of either PDCD4 or PDCD6 had no effect on HDAC3 histone deacetylase activity (Supplementary Fig. 4). Conversely, PDCD5 knockdown inhibited ET-induced reduction of HDAC3 activity, indicating that PDCD5 selectively mediates ET-induced effects on HDAC3 (Fig. 2b).


Programmed cell death 5 mediates HDAC3 decay to promote genotoxic stress response.

Choi HK, Choi Y, Park ES, Park SY, Lee SH, Seo J, Jeong MH, Jeong JW, Jeong JH, Lee PC, Choi KC, Yoon HG - Nat Commun (2015)

PDCD5 stabilizes p53 by inducing dissociation of the HDAC3–p53 complex and concomitantly triggering cytosolic cleavage of HDAC3.(a) Overexpression of PDCD5 reduces the activity of HDAC3. HCT-116 cells were transfected with the indicated plasmids. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and then HDAC3 activity was measured. Error bars, s.d. (n=3). *P<0.05. (b) Depletion of PDCD5 abolishes the ET-induced reduction of HDAC3 activity. Cells were transfected with the indicated siRNAs. Cells were treated with ET (100 μM, 12 h) or STS (1 μM, 12 h) and assayed for HDAC3 activity. Error bars, s.d. (n=3). *P<0.05. (c) Knockdown of PDCD5 abrogates the association of HDAC3 with active caspase-3 in response to ET treatment. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and subsequently immunoblotted with the indicated antibodies. Arrow indicates cleaved HDAC3. (d) Knockdown of PDCD5 prevents cytoplasmic cleavage of HDAC3 in response to ET treatment. Following cell fractionation, fractions were immunoblotted with the indicated antibodies. (e) Overexpression of PDCD5 enhances ET-induced cytoplasmic translocation of HDAC3. Immunofluorescence analysis was performed as described in the Supplementary Experimental Procedures Section. Representative images of three independent experiments are shown. (f) PDCD5 knockdown diminishes the ET-induced dissociation of HDAC3 from p53. Indicated shRNA-expressing HCT-116 cells were treated with ET. Whole-cell lysates were analysed by western blotting with the indicated antibodies. (g) Overexpression of PDCD5 dissociates HDAC3 from p53. Cells were transfected with PDCD5 plasmids. Immunoprecipitation and immunoblotting analyses were performed with the indicated antibodies. (h,i) PDCD5 increases p53 acetylation and stability via mediating HDAC3 cleavage. Cells were treated with ET (h) or cycloheximide (i) and subsequently immunoblotted with the indicated antibodies. Scale bar, 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4490383&req=5

f2: PDCD5 stabilizes p53 by inducing dissociation of the HDAC3–p53 complex and concomitantly triggering cytosolic cleavage of HDAC3.(a) Overexpression of PDCD5 reduces the activity of HDAC3. HCT-116 cells were transfected with the indicated plasmids. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and then HDAC3 activity was measured. Error bars, s.d. (n=3). *P<0.05. (b) Depletion of PDCD5 abolishes the ET-induced reduction of HDAC3 activity. Cells were transfected with the indicated siRNAs. Cells were treated with ET (100 μM, 12 h) or STS (1 μM, 12 h) and assayed for HDAC3 activity. Error bars, s.d. (n=3). *P<0.05. (c) Knockdown of PDCD5 abrogates the association of HDAC3 with active caspase-3 in response to ET treatment. Whole-cell lysates were immunoprecipitated with anti-HDAC3 antibody, and subsequently immunoblotted with the indicated antibodies. Arrow indicates cleaved HDAC3. (d) Knockdown of PDCD5 prevents cytoplasmic cleavage of HDAC3 in response to ET treatment. Following cell fractionation, fractions were immunoblotted with the indicated antibodies. (e) Overexpression of PDCD5 enhances ET-induced cytoplasmic translocation of HDAC3. Immunofluorescence analysis was performed as described in the Supplementary Experimental Procedures Section. Representative images of three independent experiments are shown. (f) PDCD5 knockdown diminishes the ET-induced dissociation of HDAC3 from p53. Indicated shRNA-expressing HCT-116 cells were treated with ET. Whole-cell lysates were analysed by western blotting with the indicated antibodies. (g) Overexpression of PDCD5 dissociates HDAC3 from p53. Cells were transfected with PDCD5 plasmids. Immunoprecipitation and immunoblotting analyses were performed with the indicated antibodies. (h,i) PDCD5 increases p53 acetylation and stability via mediating HDAC3 cleavage. Cells were treated with ET (h) or cycloheximide (i) and subsequently immunoblotted with the indicated antibodies. Scale bar, 10 μm.
Mentions: Given results suggesting PDCD5 is involved in caspase-3-dependent HDAC3 cleavage, we next investigated the role of PDCD5 in regulation of HDAC3 function. Because HDAC activity is known to be a critical function of HDAC3 (ref. 26), we first assessed the change in HDAC3 activity after either overexpression or depletion of PDCD5 in cells. An increase in wild-type PDCD5 expression, but not mutant PDCD5L6R, resulted in a decrease of HDAC3 histone deacetylase activity (Fig. 2a). However, overexpression of either PDCD4 or PDCD6 had no effect on HDAC3 histone deacetylase activity (Supplementary Fig. 4). Conversely, PDCD5 knockdown inhibited ET-induced reduction of HDAC3 activity, indicating that PDCD5 selectively mediates ET-induced effects on HDAC3 (Fig. 2b).

Bottom Line: Restoration of PDCD5(WT) in PDCD5(-/-) MEFs restores ET-induced HDAC3 cleavage.Reduction of both PDCD5 and p53, but not reduction of either protein alone, significantly enhances in vivo tumorigenicity of AGS gastric cancer cells and correlates with poor prognosis in gastric cancer patients.Our results define a mechanism for p53 activation via PDCD5-dependent HDAC3 decay under genotoxic stress conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul 120-752, Korea.

ABSTRACT
The inhibition of p53 activity by histone deacetylase 3 (HDAC3) has been reported, but the precise molecular mechanism is unknown. Here we show that programmed cell death 5 (PDCD5) selectively mediates HDAC3 dissociation from p53, which induces HDAC3 cleavage and ubiquitin-dependent proteasomal degradation. Casein kinase 2 alpha phosphorylates PDCD5 at Ser-119 to enhance its stability and importin 13-mediated nuclear translocation of PDCD5. Genetic deletion of PDCD5 abrogates etoposide (ET)-induced p53 stabilization and HDAC3 cleavage, indicating an essential role of PDCD5 in p53 activation. Restoration of PDCD5(WT) in PDCD5(-/-) MEFs restores ET-induced HDAC3 cleavage. Reduction of both PDCD5 and p53, but not reduction of either protein alone, significantly enhances in vivo tumorigenicity of AGS gastric cancer cells and correlates with poor prognosis in gastric cancer patients. Our results define a mechanism for p53 activation via PDCD5-dependent HDAC3 decay under genotoxic stress conditions.

No MeSH data available.


Related in: MedlinePlus