title: Histone deacetylase turnover and recovery in sulforaphane-treated colon cancer cells: competing actions of 14-3-3 and Pin1 in HDAC3/SMRT corepressor complex dissociation/reassembly.
abstract: BACKGROUND
  Histone deacetylase (HDAC) inhibitors are currently undergoing clinical evaluation as anti-cancer agents. Dietary constituents share certain properties of HDAC inhibitor drugs, including the ability to induce global histone acetylation, turn-on epigenetically-silenced genes, and trigger cell cycle arrest, apoptosis, or differentiation in cancer cells. One such example is sulforaphane (SFN), an isothiocyanate derived from the glucosinolate precursor glucoraphanin, which is abundant in broccoli. Here, we examined the time-course and reversibility of SFN-induced HDAC changes in human colon cancer cells.
  
  
  RESULTS
  Cells underwent progressive G2/M arrest over the period 6-72 h after SFN treatment, during which time HDAC activity increased in the vehicle-treated controls but not in SFN-treated cells. There was a time-dependent loss of class I and selected class II HDAC proteins, with HDAC3 depletion detected ahead of other HDACs. Mechanism studies revealed no apparent effect of calpain, proteasome, protease or caspase inhibitors, but HDAC3 was rescued by cycloheximide or actinomycin D treatment. Among the protein partners implicated in the HDAC3 turnover mechanism, silencing mediator for retinoid and thyroid hormone receptors (SMRT) was phosphorylated in the nucleus within 6 h of SFN treatment, as was HDAC3 itself. Co-immunoprecipitation assays revealed SFN-induced dissociation of HDAC3/SMRT complexes coinciding with increased binding of HDAC3 to 14-3-3 and peptidyl-prolyl cis/trans isomerase 1 (Pin1). Pin1 knockdown blocked the SFN-induced loss of HDAC3. Finally, SFN treatment for 6 or 24 h followed by SFN removal from the culture media led to complete recovery of HDAC activity and HDAC protein expression, during which time cells were released from G2/M arrest.
  
  
  CONCLUSIONS
  The current investigation supports a model in which protein kinase CK2 phosphorylates SMRT and HDAC3 in the nucleus, resulting in dissociation of the corepressor complex and enhanced binding of HDAC3 to 14-3-3 or Pin1. In the cytoplasm, release of HDAC3 from 14-3-3 followed by nuclear import is postulated to compete with a Pin1 pathway that directs HDAC3 for degradation. The latter pathway predominates in colon cancer cells exposed continuously to SFN, whereas the former pathway is likely to be favored when SFN has been removed within 24 h, allowing recovery from cell cycle arrest.
authors.affiliation: Linus Pauling Institute, Oregon State University, Corvallis, OR 97331-6512, USA.
authors.forename: Praveen
authors.initials: P
authors.lastname: Rajendran
authors.forename: Barbara
authors.initials: B
authors.lastname: Delage
authors.forename: W Mohaiza
authors.initials: WM
authors.lastname: Dashwood
authors.forename: Tian-Wei
authors.initials: TW
authors.lastname: Yu
authors.forename: Bradyn
authors.initials: B
authors.lastname: Wuth
authors.forename: David E
authors.initials: DE
authors.lastname: Williams
authors.forename: Emily
authors.initials: E
authors.lastname: Ho
authors.forename: Roderick H
authors.initials: RH
authors.lastname: Dashwood
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism*; D000107:Acetylation / Q000187:drug effects; D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000201:enzymology / Q000503:physiopathology*; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D045325:HCT116 Cells; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006655:Histone Deacetylases / Q000378:metabolism*; D006801:Humans; D017879:Isothiocyanates; D008247:Lysosomes / Q000378:metabolism; D008954:Models, Biological; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D056985:Nuclear Receptor Co-Repressor 2 / Q000378:metabolism*; D019696:Peptidylprolyl Isomerase / Q000378:metabolism*; D061988:Proteasome Inhibitors; D011485:Protein Binding / Q000502:physiology; D013454:Sulfoxides; D013861:Thiocyanates / Q000494:pharmacology*
chemical_list: D048948:14-3-3 Proteins; D016588:Anticarcinogenic Agents; D056572:Histone Deacetylase Inhibitors; D017879:Isothiocyanates; C539130:NCOR2 protein, human; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D056985:Nuclear Receptor Co-Repressor 2; D061988:Proteasome Inhibitors; D013454:Sulfoxides; D013861:Thiocyanates; D003609:Dactinomycin; D003513:Cycloheximide; D053148:Caspase 3; D006655:Histone Deacetylases; C000606846:PIN1 protein, human; D019696:Peptidylprolyl Isomerase; C016766:sulforaphane