Single-molecule force spectroscopy reveals a highly compliant helical folding for the 30-nm chromatin fiber

Abstract

The compaction of eukaryotic DNA into chromatin has been implicated in the regulation of all DNA processes. To unravel the higher-order folding of chromatin, we used magnetic tweezers and probed the mechanical properties of single 197-bp repeat length arrays of 25 nucleosomes. At forces up to 4 pN, the 30-nm fiber stretches like a Hookian spring, resulting in a three-fold extension. Together with a high nucleosome-nucleosome stacking energy, this points to a solenoid as the underlying topology of the 30-nm fiber. Unexpectedly, linker histones do not affect the length or stiffness of the fiber but stabilize its folding. Fibers with a nucleosome repeat length of 167 bp are stiffer, consistent with a two-start helical arrangement. The observed high compliance causes extensive thermal breathing, which forms a physical basis for the balance between DNA condensation and accessibility.

The energy components of stacked chromatin layers explain the morphology, dimensions and mechanical properties of metaphase chromosomes

“…Young modulus Y (kPa) corresponding to the elastic extension of the chromosomes considered in this table were obtained from [30 -32,34,35]. The total energy due to nucleosome-nucleosome interactions E nn (pJ) was calculated from the number of nucleosomes N n in each chromosome (N n % number of bp per chromosome/200 bp per nucleosome) and considering previous experimental [21,22] and modelling [25 -27] studies showing that the energy 1 nn of a single internucleosome interaction is between 3.4 and 14 k B T (E nn ¼ N n 1 nn ; see equation (2.5) contiguous scaffold formed by non-histone proteins. Nevertheless, different evidences indicated that condensins and topoisomerase II, which are located in the chromatid axis, may play a role in the functional organization of chromosomes [64].…”

“…Thus, it is very likely that the energy 1 nn that stabilizes the stacking of the successive layers in chromosomes is essentially due to face-to-face interactions between nucleosomes. The energy per nucleosome in internucleosome interactions has been obtained experimentally [21,22] and in modelling studies [25 -27] in several laboratories (see Introduction); taking into account all the values obtained in these studies, it is reasonable to consider that 1 nn % 3.4 -14 k B T. On the other hand, the easy sliding between layers in chromatin plates observed in electron microscopy experiments [15,16], suggested that the forces holding the chromatin filament within a layer are higher that the interactions between adjacent layers. This is consistent with the relatively large the Young modulus [15] and the high mechanical resistance [17] found for chromatin plates in AFM experiments in aqueous media, and indicates that the energy 1 wl stabilizing the structure of layers is higher than 1 nn .…”

“…The force-extension curves corresponding to the unfolding and refolding of the fibres indicated that this process is reversible and allowed the calculation of the energy corresponding to the disruption of the attractive interactions between nucleosomes in folded fibres. The energy required for breaking a single nucleosome-nucleosome interaction is between 3.4 and 14 k B T [21,22] (%3.6 Â 10 220 J considering that k B T ¼ 4.1 Â 10 221 J; k B is the Boltzmann constant and T corresponds to the room temperature). These experimental results are compatible with the internucleosome interaction energy obtained in modelling studies of chromatin fibres having different degrees of nucleosome stacking [25][26][27].…”

“…The elongated shape of chromatids can be explained if the destabilizing surface potential is higher in the telomeres (approx. 0.16 mJ m 22 ) than in the lateral surface (approx. 0.012 mJ m 22 ).…”

“…0.16 mJ m 22 ) than in the lateral surface (approx. 0.012 mJ m 22 ). The results obtained by other authors in experimental studies of chromosome mechanics have been used to test the proposed supramolecular structure.…”