In vivo DNA topology and conformations of chromatin

Abstract

In contrast to our approximated view of the topology and conformations of naked DNA molecules in vitro, we can hardly picture the topology and spatial configurations of DNA in vivo. Chromatin architecture and its enzymatic activities determine how twist and writhe deformations of DNA are constrained or unconstrained, and how DNA supercoiling energy is generated, buffered or dissipated at each genomic site. Understanding the topology of nucleosomal DNA and nucleosomal fibers is therefore a fundamental prerequisite to evaluate the DNA topology outcomes of more complex chromatin structures. In this regard, we have revisited the "linking number paradox" of nucleosomal DNA, which states that the 1.7 lefthanded turns of DNA around a histone octamer results in the apparent stabilization of only one negative DNA supercoil (∆Lk ≈ -1). In order to assess how nucleosomal fibers are packaged in vivo, we have also conducted a first analysis of the probability of DNA knot formation in eukaryotic chromatin. We found that steady state fractions of trefoils and more complex knot species are maintained by topoisomerase II activity. We uncovered also that the knotting probability of intracellular DNA does not scale proportionally to chromatin length. On the basis of our experimental observations, we inferred novel traits of the nucleosomal DNA topology and the chromatin architecture in vivo.