Ukr.Biochem.J. 2014; Volume 86, Issue 4, Jul-Aug, pp. 110-118

doi: http://dx.doi.org/10.15407/ubj86.04.110

Protein intercalation in DNA as one of main modes of fixation of the most stable chromatin loop domains

М. I. Chopei, K. S. Afanasieva, А. V. Sivolob

Taras Shevchenko National University of Kyiv, Ukraine;
e-mail: aphon@ukr.net; sivolob@univ.kiev.ua

The main mechanism of DNA track formation during comet assay of nucleoids, obtained after removal of cell membranes and most of proteins, is the extension to anode of negatively supercoiled DNA loops attached to proteins, remaining in nucleoid after lysis treatment. The composition of these residual protein structures and the nature of their strong interaction with the loop ends remain poorly studied. In this work we investigated the influence of chloroquine intercalation and denaturation of nucleoid proteins on the efficiency of electrophoretic track formation during comet assay. The results obtained suggest that even gentle protein denaturation is sufficient to reduce considerably the effectiveness of the DNA loop migration due to an increase in the loops size. The same effect was observed under local DNA unwinding upon chloroquine intercalation around the sites of the attachment of DNA to proteins. The topological interaction (protein intercalation into the double helix) between DNA loop ends and nucleoid proteins is discussed.

Keywords: , , , ,


References:

  1. Van Bortle K, Corces VG. Nuclear organization and genome function. Annu Rev Cell Dev Biol. 2012;28:163-87. Review. PubMed, PubMedCentral, CrossRef
  2. Kadauke S, Blobel GA. Chromatin loops in gene regulation. Biochim Biophys Acta. 2009 Jan;1789(1):17-25. Review. PubMed, PubMedCentral, CrossRef
  3. Cook PR. A model for all genomes: the role of transcription factories. J Mol Biol. 2010 Jan 8;395(1):1-10. Review. PubMed, CrossRef
  4. Cremer T, Cremer C. Chromosome territories, nuclear architecture and gene regulation in mammalian cells. Nat Rev Genet. 2001 Apr;2(4):292-301. Review. PubMed
  5. Malyavantham KS, Bhattacharya S, Berezney R. The architecture of functional neighborhoods within the mammalian cell nucleus. Adv Enzyme Regul. 2010;50(1):126-34. Review.  PubMed, PubMedCentral, CrossRef
  6. Bohn M, Heermann DW. Diffusion-driven looping provides a consistent framework for chromatin organization. PLoS One. 2010 Aug 25;5(8):e12218. PubMed, PubMedCentral, CrossRef
  7. Iarovaia OV, Lagarkova MA, Razin SV. The specificity of human lymphocyte nucleolar DNA long-range fragmentation by endogenous topoisomerase II and exogenous Bal 31 nuclease depends on cell proliferation status. Biochemistry. 1995 Mar 28;34(12):4133-8. PubMed, CrossRef
  8. Vassetzky Y, Hair A, Méchali M. Rearrangement of chromatin domains during development in Xenopus. Genes Dev. 2000 Jun 15;14(12):1541-52. PubMed, PubMedCentral
  9. Hancock R. Internal organisation of the nucleus: assembly of compartments by macromolecular crowding and the nuclear matrix model. Biol Cell. 2004 Oct;96(8):595-601. Review. PubMed, CrossRef
  10. Pederson T. Half a century of “the nuclear matrix”. Mol Biol Cell. 2000 Mar;11(3):799-805. Review. PubMed, PubMedCentral, CrossRef
  11. Hancock R. A new look at the nuclear matrix. Chromosoma. 2000 Jul;109(4):219-25. PubMed, CrossRef
  12. Cook PR, Brazell IA. Supercoils in human DNA. J Cell Sci. 1975 Nov;19(2):261-79. PubMed
  13. Olive PL. The comet assay. An overview of techniques. Methods Mol Biol. 2002;203:179-94. Review. PubMed, CrossRef
  14. Collins AR, Oscoz AA, Brunborg G, Gaivão I, Giovannelli L, Kruszewski M, Smith CC, Stetina R. The comet assay: topical issues. Mutagenesis. 2008 May;23(3):143-51. Review. PubMed, CrossRef
  15. Afanas’eva KS, Zazhitskaia MO, Sivolob AV. Mechanisms of DNA exit during neutral and alkaline comet assay. Tsitol Genet. 2009 Nov-Dec;43(6):3-7. Russian. PubMed
  16. Afanasieva K, Zazhytska M, Sivolob A. Kinetics of comet formation in single-cell gel electrophoresis: loops and fragments. Electrophoresis. 2010 Jan;31(3):512-9. PubMed, CrossRef
  17. Zazhytska M, Afanasieva K, Chopei M, Vikhreva M, Sivolob A. Influence of chloroquine on kinetics of single-cell gel electrophoresis. Biopolym  Cell. 2012;28(4):292–297. CrossRef
  18.  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 Dec;1833(12):3237-44. PubMed, CrossRef
  19. Jones RL, Lanier AC, Keel RA, Wilson WD. The effect of ionic strength on DNA-ligand unwinding angles for acridine and quinoline derivatives. Nucleic Acids Res. 1980 Apr 11;8(7):1613-24. PubMed, PubMedCentral

Creative CommonsThis work is licensed under a Creative Commons Attribution 4.0 International License.