Despite the key role of the linker histone H1 in chromatin structure and dynamics, its location and interactions with nucleosomal DNA have not been elucidated. In this work we have used a combination of electron cryomicroscopy, hydroxyl radical footprinting, and nanoscale modeling to analyze the structure of precisely positioned mono-, di-, and trinucleosomes containing physiologically assembled full-length histone H1 or truncated mutants of this protein.Single-base resolution •OH footprinting shows that the globular domain of histone H1 (GH1) interacts with the DNA minor groove located at the center of the nucleosome and contacts a 10-bp region of DNA localized symmetrically with respect to the nucleosomal dyad. In addition, GH1 interacts with and organizes about one helical turn of DNA in each linker region of the nucleosome. We also find that a seven amino acid residue region (121-127) in the COOH terminus of histone H1 was required for the formation of the stem structure of the linker DNA. A molecular model on the basis of these data and coarse-grain DNA mechanics provides novel insights on how the different domains of H1 interact with the nucleosome and predicts a specific H1-mediated stem structure within linker DNA.nucleosome structure | chromatin higher order structure T he nucleosome is the fundamental repeating unit of chromatin in the nucleus of eukaryotic cells. The composition and the basic organization of the nucleosome is well established, and the structure of the nucleosomal core particle (NCP) has been described with nearly atomic precision by X-ray diffraction (1). However, similar information for the structure of a complete nucleosome, i.e., the NCP with associated linker DNA segments and a linker histone, is still lacking. Electron microscopy and electron cryomicroscopy (ECM) imaging have provided a relatively low-resolution picture of the complete nucleosome, both native (2) and reconstituted (3). However, important features of the structure remain obscure.Linker histones are typically ∼200 aa in length with a rather short nonstructured N terminus, followed by a ∼70-80 aa structured ("globular") domain, and a ∼100 aa long apparently unstructured C terminal domain, highly enriched in lysines. The globular domain of the linker histone appears to be internally located in the 30-nm chromatin fiber (4, 5), but its exact position within the nucleosome remains a subject of debate (for review, see ref. 6). A second question not yet resolved concerns the interactions and location of the linker histone C terminus. These issues have their origin in difficulties related to the preparation of well-defined nucleosomal samples. Indeed, direct binding of linker histone to nucleosomes in vitro is inefficient and complicated by the formation of large aggregates because of the nonspecific association of linker histones with DNA (7, 8).The situation can be considerably improved by using chaperones for linker histone deposition in vitro, a mechanism that is likely used in vivo (9). It was recently shown that NAP...

DNA torsional stress is generated by virtually all biomolecular processes involving the double helix, in particular transcription where a significant level of stress propagates over several kilobases. If another promoter is located in this range, this stress may strongly modify its opening properties, and hence facilitate or hinder its transcription. This mechanism implies that transcribed genes distant of a few kilobases are not independent, but coupled by torsional stress, an effect for which we propose the first quantitative and systematic model. In contrast to previously proposed mechanisms of transcriptional interference, the suggested coupling is not mediated by the transcription machineries, but results from the universal mechanical features of the double-helix. The model shows that the effect likely affects prokaryotes as well as eukaryotes, but with different consequences owing to their different basal levels of torsion. It also depends crucially on the relative orientation of the genes, enhancing the expression of eukaryotic divergent pairs while reducing that of prokaryotic convergent ones. To test the in vivo influence of the torsional coupling, we analyze the expression of isolated gene pairs in the Drosophila melanogaster genome. Their orientation and distance dependence is fully consistent with the model, suggesting that torsional gene coupling may constitute a widespread mechanism of (co)regulation in eukaryotes.