title: Concordant and opposite roles of DNA-PK and the "facilitator of chromatin transcription" (FACT) in DNA repair, apoptosis and necrosis after cisplatin.
abstract: BACKGROUND
  Platinum-containing chemotherapy produces specific DNA damage and is used to treat several human solid tumors. Tumors initially sensitive to platinum-based drugs frequently become resistant. Inhibition of DNA repair is a potential strategy to enhance cisplatin effectiveness. After cisplatin treatment, a balance between repair and apoptosis determines whether cancer cells proliferate or die. DNA-dependent protein kinase (DNA-PK) binds to DNA double strand breaks (DSBs) through its Ku subunits and initiates non-homologous end joining. Inhibition of DNA-PK sensitizes cancer cells to cisplatin killing. The goal of this study is to elucidate the mechanism underlying the effects of DNA-PK on cisplatin sensitivity.
  
  
  RESULTS
  Silencing the expression of the catalytic subunit of DNA-PK (DNA-PKcs) increased sensitivity to cisplatin and decreased the appearance of γH2AX after cisplatin treatment. We purified DNA-PK by its Ku86 subunit and identified interactors by tandem mass spectrometry before and after cisplatin treatment. The structure specific recognition protein 1 (SSRP1), Spt16 and γH2AX appeared in the Ku86 complex 5 hours after cisplatin treatment. SSRP1 and Spt16 form the facilitator of chromatin transcription (FACT). The cisplatin-induced association of FACT with Ku86 and γH2AX was abrogated by DNase treatment. In living cells, SSRP1 and Ku86 were recruited at sites of DSBs induced by laser beams. Silencing SSRP1 expression increased sensitivity to cisplatin and decreased γH2AX appearance. However, while silencing SSRP1 in cisplatin-treated cells increased both apoptosis and necrosis, DNA-PKcs silencing, in contrast, favored necrosis over apoptosis.
  
  
  CONCLUSIONS
  DNA-PK and FACT both play roles in DNA repair. Therefore both are putative targets for therapeutic inhibition. Since DNA-PK regulates apoptosis, silencing DNA-PKcs redirects cells treated with cisplatin toward necrosis. Silencing FACT however, allows both apoptosis and necrosis. Targeting DNA repair in cancer patients may have different therapeutic effects depending upon the roles played by factors targeted.
authors.affiliation: Department of Cancer Biology, Dana-Farber Cancer Institute, 450 Brookline Avenue, Boston, Massachusetts 02215, USA.
authors.forename: Janna
authors.initials: J
authors.lastname: Sand-Dejmek
authors.forename: Guillaume
authors.initials: G
authors.lastname: Adelmant
authors.forename: Bijan
authors.initials: B
authors.lastname: Sobhian
authors.forename: Anne S
authors.initials: AS
authors.lastname: Calkins
authors.forename: Jarrod
authors.initials: J
authors.lastname: Marto
authors.forename: Dirk J
authors.initials: DJ
authors.lastname: Iglehart
authors.forename: Jean-Bernard
authors.initials: JB
authors.lastname: Lazaro
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000494:pharmacology*; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects; D004249:DNA Damage / Q000187:drug effects / Q000235:genetics; D004260:DNA Repair / Q000187:drug effects* / Q000235:genetics; D051747:DNA-Activated Protein Kinase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D004353:Drug Evaluation, Preclinical; D057809:HEK293 Cells; D006367:HeLa Cells; D006609:High Mobility Group Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D009336:Necrosis / Q000139:chemically induced / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D035602:Transcriptional Elongation Factors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; C073537:SSRP1 protein, human; D035602:Transcriptional Elongation Factors; D051747:DNA-Activated Protein Kinase; D002945:Cisplatin