ИСТИНА

Introduction: RUNX1(AML1)-associated chromosomal translocations is a marked feature of “treatment-related” leukemias caused by DNA-topoisomerase inhibitors (e.g. etoposide) widely used in anti-cancer therapy. This pathogenic process occurs when erroneous repair of etoposide-induced double-stranded DNA breaks within RUNX1 gene leads to the chromosomal rearrangements. The molecular mechanisms of such events may be unraveled by using the fluorescent microscopy accompanied by data processing. Methods: We treated cultured human T-lymphocytes (Jurkat) with etoposide and inhibitors of Rad51 and Mre11 (B02 and mirin, correspondingly), which participate in double-stranded DNA breaks repair and fixed the cells with paraformaldehyde. 3D-FISH was performed with using RUNX1 dual color break-apart probe and whole chromosome-21 painting probe. Confocal images obtained were processed by means of specially designed computer software allowing to detect the signals and to measure the distances between them. Results: Etoposide promotes the double-stranded DNA breaks within RUNX1 with measurable frequency. Broken alleles RUNX1 alleles used to be located outside the corresponding chromosome territory more often than non-broken ones. Both inhibitors, mirin (Mre11 inhibitor) and B02 (Rad51 inhibitor) does not affect this phenomenon and does not lead to a significant decrease in DNA breakage efficiency. Conclusions: Data obtained by wide-scale observation of chromatin dynamics by meas of 3D-FISH and computer analysis indicate that Rad51 and Mre11 does not affect the mobility of broken and non-broken alleles of RUNX1 after induction of double-stranded DNA breaks by etoposide. This study was partially supported by the Russian Foundation of Basic Research (grants 14-04-93105_CNRS_а, 15-54-16007_CNRS_а).