Phosphorylated Sp1 is the regulator of DNA-PKcs and DNA ligase IV transcription of daunorubicin-resistant leukemia cell lines

Yayoi Nishida, Naoki Mizutani, Minami Inoue, Yukari Omori, Keiko Tamiya-Koizumi, Akira Takagi, Tetsuhito Kojima, Motoshi Suzuki, Yoshinori Nozawa, Yosuke Minami, Kazunori Ohnishi, Tomoki Naoe, Takashi Murate

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14 Citations (Scopus)

Abstract

Multidrug resistance (MDR) is a serious problem faced in the treatment of malignant tumors. In this study, we characterized the expression of non-homologous DNA end joining (NHEJ) components, a major DNA double strand break (DSB) repair mechanism in mammals, in K562 cell and its daunorubicin (DNR)-resistant subclone (K562/DNR). K562/DNR overexpressed major enzymes of NHEJ, DNA-PKcs and DNA ligase IV, and K562/DNR repaired DSB more rapidly than K562 after DNA damage by neocarzinostatin (MDR1-independent radiation-mimetic). Overexpressed DNA-PKcs and DNA ligase IV were also observed in DNR-resistant HL60 (HL60/DNR) cells as compared with parental HL60 cells. Expression level of DNA-PKcs mRNA paralleled its protein level, and the promoter activity of DNA-PKcs of K562/DNR was higher than that of K562, and the 5'-region between - 49. bp and the first exon was important for its activity. Because this region is GC-rich, we tried to suppress Sp1 family transcription factor using mithramycin A (MMA), a specific Sp1 family inhibitor, and siRNAs for Sp1 and Sp3. Both MMA and siRNAs suppressed DNA-PKcs expression. Higher serine-phosphorylated Sp1 but not total Sp1 of both K562/DNR and HL60/DNR was observed compared with their parental K562 and HL60 cells. DNA ligase IV expression of K562/DNR was also suppressed significantly with Sp1 family protein inhibition. EMSA and ChIP assay confirmed higher binding of Sp1 and Sp3 with DNA-PKcs 5'-promoter region of DNA-PKcs of K562/DNR than that of K562. Thus, the Sp1 family transcription factor affects important NHEJ component expressions in anti-cancer drug-resistant malignant cells, leading to the more aggressive MDR phenotype.

Original languageEnglish
Pages (from-to)265-274
Number of pages10
JournalBiochimica et Biophysica Acta - Gene Regulatory Mechanisms
Volume1839
Issue number4
DOIs
Publication statusPublished - 04-2014

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics

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    Nishida, Y., Mizutani, N., Inoue, M., Omori, Y., Tamiya-Koizumi, K., Takagi, A., Kojima, T., Suzuki, M., Nozawa, Y., Minami, Y., Ohnishi, K., Naoe, T., & Murate, T. (2014). Phosphorylated Sp1 is the regulator of DNA-PKcs and DNA ligase IV transcription of daunorubicin-resistant leukemia cell lines. Biochimica et Biophysica Acta - Gene Regulatory Mechanisms, 1839(4), 265-274. https://doi.org/10.1016/j.bbagrm.2014.02.004