Changes in loop organization of chromatin at different stages of lymphocyte activation

  • K. S. Afanasieva Taras Shevchenko National University of Kyiv, Ukraine, 01601, Kyiv, Volodimyrska str., 64/13
  • O. V. Lozovik Taras Shevchenko National University of Kyiv, Ukraine, 01601, Kyiv, Volodimyrska str., 64/13
  • V. V. Olefirenko Taras Shevchenko National University of Kyiv, Ukraine, 01601, Kyiv, Volodimyrska str., 64/13
  • A. V. Sivolob Taras Shevchenko National University of Kyiv, Ukraine, 01601, Kyiv, Volodimyrska str., 64/13

Abstract

Aim. Aim was to investigate possible changes in the DNA loop domain organization upon activation of human lymphocytes. The rational for this task is the knowledge that the chromatin looping plays an important role in transcription regulation and thus may vary depending on cell functional state. Methods. The kinetics of DNA loop migration during single cell gel electrophoresis (the comet assay) was studied for nucleoids obtained from human lymphocytes and lymphoblasts activated to proliferation by interleukin 2. Results. Three part of DNA were observed in nucleoids: DNA on the nucleoid surface, loops up to ~150 kb inside the nucleoid, and larger loops that cannot migrate. An essential redistribution of the loop domains between the inside and surface fractions occurs upon activation (at G1 phase). Later on (at the end of S phase) the inside fraction becomes lower in favor of the large loops. Conclusions. Changes in the cell functional state are accompanied by large-scale changes in the loop domain organization that can be detected by the comet assay.
Keywords: DNA loops, nucleoid, comet assay, lymphocytes, lymphoblasts.

References

Dekker J., Marti-Renom M.A., Mirny L.A. Exploring the three-dimensional organization of genomes: interpreting chromatin interaction data. Nat. Rev. Genet. 2013. V. 14. P. 390–403. doi: 10.1038/nrg3454.

Dekker J., Mirny L. The 3D genome as moderator of chromosomal communication. Cell. 2016. V. 164. P. 1110–1121. doi: 10.1016/j.cell.2016.02.007.

Dixon J.R., Gorkin D.U., Ren B. Chromatin domains: the unit of chromosome organization. Mol. Cell. 2016. V. 62. P. 668–680. doi: 10.1016/j.molcel.2016.05.018.

Rao S.S.P., Huntley M.H., Durand N.C., Stamenova E.K., Bochkov I.D., Robinson J.T., Sanborn A.L., Machol I., Omer A.D., Lander E.S., Aiden E.L. A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell. 2014. V. 159. P. 1665–1680. doi: 10.1016/j.cell.2014.11.021.

Dixon J.R., Selvaraj S., Yue F., Kim A., Li Y., Shen Y., Hu M., Liu J.S., Ren B. Topological domains in mammalian genomes identified by analysis of chromatin interactions. Nature. 2012. V. 485. P. 376–380. doi: 10.1038/nature11082.

Cook P.R. A model for all genomes: the role of transcription factories. J. Mol. Biol. 2010. V. 395. P. 1–10. doi: 10.1016/j.jmb.2009.10.031.

Tang Z., Luo O. J., Li X., Plewczynski D., Li G., Ruan Y. CTCF-Mediated Human 3D Genome Architecture Reveals Chromatin Topology for Transcription. Cell. 2015. V. 163. P. 1611–1627. doi: 10.1016/j.cell.2015.11.024.

Afanasieva K., Zazhytska M., Sivolob A. Kinetics of comet formation in single-cell gel electrophoresis: loops and fragments. Electrophoresis. 2010. V. 31. P. 512–519. doi: 10.1002/elps.200900421.

Afanasieva K., Chopei M., Zazhytska M. Vikhreva M., Sivolob A. DNA loop domain organization as revealed by single-cell gel electrophoresis. Biochim. Biophys. Acta. 2013. V. 1833. P. 3237–3244. doi: 10.1016/j.bbamcr.2013.09.021.

Shaposhnikov S.A, Salenko V.B., Brunborg G., Nygren J., Collins A.R. Single-cell gel electrophoresis (the comet assay): loops or fragments. Electrophoresis. 2008. V. 29. P. 3005–3012. doi: 10.1002/elps.200700921.

Collins A.R., Oscoz A.A., Brunborg G., Gaivao I., Giovannelli L., Kruszewski M., Stetina R., Smith C.C. The comet assay: topical issues. Mutagenesis. 2008. V. 23. P. 143–151. doi: 10.1093/mutage/gem051.

Afanasieva K., Chopei M., Lozovik A., Semenova A., Lukash L., Sivolob A. DNA loop domain organization in nucleoids from cells of different types. Biochem. Biophys. Res. Commun. 2017. V. 483. P. 142–146. doi: 10.1016/j.bbrc.2016.12.177.

Zazhytska M., Afanasieva K., Chopei M., Vikhreva M., Sivolob A. Influence of chloroquine on kinetics of single-cell gel electrophoresis. Biopolym.Cell. 2012. V. 28. P. 292–297. doi: 10.7124/bc.000062.

Mzali R., Seguin L., Liot C., Auger A., Pacaud P., Loirand G., Thibault C., Pierre J., Bertoglio J. Regulation of Rho signaling pathways in interleukin-2-stimulated human T-lymphocytes. FASEB J. 2005. V. 219. P. 1911–1933. doi: 10.1096/fj.05-4030fje.

Kaplan O., Aebersold P., Cohen J.S. Metabolism of peripheral lymphocytes, interleukin-2 activated lymphocytes and tumorinfiltrating lymphocytes from 31P NMR studies. FEBS Lett. 1989. V. 258. P. 55–58. doi: 10.1016/0014-5793(89)81614-X

Bender M.A., Prescott D.M. DNA synthesis and mitosis in cultures of human peripheral leukocytes. Exp. Cell Res. 1962. V. 27. P. 221–229. doi: 10.1016/0014-4827(62)90225-2

Bain B.J. Blood cells: a practical guide. Oxford: Blackwell Publishing, 2006. 476 p. doi: 10.1002/9780470987551