title: HDAC inhibition activates the apoptosome via Apaf1 upregulation in hepatocellular carcinoma.
abstract: BACKGROUND
  Histone deacetylation, a common hallmark in malignant tumors, strongly alters the transcription of genes involved in the control of proliferation, cell survival, differentiation and genetic stability. We have previously shown that HDAC1, HDAC2, and HDAC3 (HDAC1-3) genes encoding histone deacetylases 1-3 are upregulated in primary human hepatocellular carcinoma (HCC). The aim of this study was to characterize the functional effects of HDAC1-3 downregulation and to identify functionally important target genes of histone deacetylation in HCC.
  
  
  METHODS
  Therefore, HCC cell lines were treated with the histone deacetylase inhibitor (HDACi) trichostatin A and by siRNA-knockdown of HDAC1-3. Differentially expressed mRNAs were identified after siRNA-knockdown of HDAC1-3 using mRNA expression profiling. Findings were validated after siRNA-mediated silencing of HDAC1-3 using qRTPCR and Western blotting assays.
  
  
  RESULTS
  mRNA profiling identified apoptotic protease-activating factor 1 (Apaf1) to be significantly upregulated after HDAC inhibition (HLE siRNA#1/siRNA#2 p < 0.05, HLF siRNA#1/siRNA#2 p < 0.05). As a component of the apoptosome, a caspase-activating complex, Apaf1 plays a central role in the mitochondrial caspase activation pathway of apoptosis. Using annexin V, a significant increase in apoptosis could also be shown in HLE (siRNA #1 p = 0.0034) and HLF after siRNA against HDAC1-3 (Fig. 3a, b). In parallel, caspase-9 activity was increased after siRNA-knockdown of HDAC1-3 leading to enhanced apoptosis after HDAC inhibition (Fig. 3c, d).
  
  
  CONCLUSIONS
  The present data show that siRNA-knockdown of HDAC1-3 plays a major role in mediating apoptotic response to HDAC inhibitors through regulation of Apaf1.
authors.affiliation: Institute of Human Genetics, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany.
authors.forename: Reena
authors.initials: R
authors.lastname: Buurman
authors.affiliation: Institute of Human Genetics, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Sandbothe
authors.affiliation: Institute of Human Genetics, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Schlegelberger
authors.affiliation: Institute of Human Genetics, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany. Skawran.britta@mh-hannover.de.
authors.forename: Britta
authors.initials: B
authors.lastname: Skawran
mesh_terms: D017209:Apoptosis; D053477:Apoptotic Protease-Activating Factor 1 / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D020169:Caspases / Q000378:metabolism; D049109:Cell Proliferation; D056284:Histone Deacetylase 1 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D056464:Histone Deacetylase 2 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D006655:Histone Deacetylases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D008928:Mitochondria / Q000378:metabolism; D011379:Prognosis; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D000099186:Histone Deacetylase 3
chemical_list: D053477:Apoptotic Protease-Activating Factor 1; D020169:Caspases; D056284:Histone Deacetylase 1; D056464:Histone Deacetylase 2; D006655:Histone Deacetylases; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000099186:Histone Deacetylase 3; C506592:APAF1 protein, human; C110550:HDAC1 protein, human; C533423:HDAC2 protein, human
keywords: Acetylation; Apoptosis; Hepatocellular carcinoma; Histone deacetylases; Trichostatin A