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The effects of the HDAC inhibitor sodium butyrate on the expression of repair genes Rad51 and XRCC5 in fibroblast lines mEras-Waf1+/+ and mEras-Waf1–/–

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Abstract

Transformed mice fibroblasts with a knockout CDKN1A gene, which encodes р21/Waf1 protein (mEras-Waf1–/–cells), are characterized by a large number of single-strand DNA breaks and associated γ-H2A.X foci compared to the initial transformant line E1A+cHa-ras (mEras-Waf1+/+ cells). According to immunofluorescence and immunoblotting data, the nuclei of the cells from both studied lines mEras-Waf1+/+ and mEras-Waf1–/–contain significant amounts of Rad51 and Ku80 proteins, which participate in DNA repair, with a slight prevalence of Ku80 in CDKN1A knockout cells. Under a short-term effect of adriamycin, a DNA-damaging agent, on the cells of both lines, additional accumulation of Rad51 protein foci occurs in their nuclei. However, an HDAC inhibitor, sodium butyrate, notably decreases the contents of Rad51 and Ku80 proteins in both intact mEras-Waf1+/+ and mEras-Waf1–/–cells and cells treated with adriamycin. Results of RT-PCR and immunoblotting demonstrate that inhibiting effect of sodium butyrate (NaBut) becomes apparent at the level of Rad51 and XRCC5 gene transcription and at the level of translation of the respective repair proteins, Rad51 and Ku80. The observed suppressive effect of NaBut (HDAC inhibitor) on the components of DNA repair system can be partially explained by antiproliferative function of HDAC inhibitors. In addition, transcriptional activation of pluripotency genes Oct-4, Sox-2, and Klf4 was discovered in mEras-Waf1+/+ and mEras-Waf1–/–cells under the influence of NaBut, which implies that these genes are under negative control at the level of the chromatin structure.

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Abbreviations

RT:

reverse transcription

PAGE:

polyacrylamide gel

PCR:

polymerase chain reaction

DDR:

DNA damage response

DSB:

double-strand breaks

HDAC:

histone deacetylases

NaBut:

sodium butyrate

NHEJ:

nonhomologous end joining

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Authors and Affiliations

  1. Institute of Cytology, Russian Academy of Sciences, St. Petersburg, 194064, Russia

    A. N. Kukushkin, S. B. Svetlikova & V. A. Pospelov

Authors
  1. A. N. Kukushkin
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  2. S. B. Svetlikova
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  3. V. A. Pospelov
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Correspondence to A. N. Kukushkin.

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Original Russian Text © A.N. Kukushkin, S.B. Svetlikova, V.A. Pospelov, 2016, published in Tsitologiya, 2016, Vol. 58, No. 12, pp. 908–915.

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Kukushkin, A.N., Svetlikova, S.B. & Pospelov, V.A. The effects of the HDAC inhibitor sodium butyrate on the expression of repair genes Rad51 and XRCC5 in fibroblast lines mEras-Waf1+/+ and mEras-Waf1–/–. Cell Tiss. Biol. 11, 133–140 (2017). https://doi.org/10.1134/S1990519X1702002X

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  • DOI: https://doi.org/10.1134/S1990519X1702002X

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