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SummaryLysine acetylation of histones defines the epigenetic status of human embryonic stem cells and orchestrates DNA replication, chromosome condensation, transcription, telomeric silencing, and DNA repair. A detailed mechanistic explanation of these phenomena is impeded by the limited availability of homogeneously acetylated histones. We report a general method for the production of homogeneously and site-specifically acetylated recombinant histones by genetically encoding acetyl-lysine. We reconstitute histone octamers, nucleosomes, and nucleosomal arrays bearing defined acetylated lysine residues. With these designer nucleosomes, we demonstrate that, in contrast to the prevailing dogma, acetylation of H3 K56 does not directly affect the compaction of chromatin and has modest effects on remodeling by SWI/SNF and RSC. Single-molecule FRET experiments reveal that H3 K56 acetylation increases DNA breathing 7-fold. Our results provide a molecular and mechanistic underpinning for cellular phenomena that have been linked with K56 acetylation.

Summary Currently there are no approved vaccines or specific therapies to prevent or treat Zika virus (ZIKV) infection. We interrogated a library of FDA-approved drugs for their ability to block infection of human HuH-7 cells by a newly isolated ZIKV strain (ZIKV MEX_I_7). More than 20 out of 774 tested compounds decreased ZIKV infection in our in vitro screening assay. Selected compounds were further validated for inhibition of ZIKV infection in human cervical, placental and neural stem cell lines, as well as primary human amnion cells. Established anti-flaviviral drugs (e.g., bortezomib and mycophenolic acid) and others that had no previously known anti-viral activity (e.g., daptomycin) were identified as inhibitors of ZIKV infection. Several drugs reduced ZIKV infection across multiple cell types. This study identifies drugs that could be tested in clinical studies of ZIKV infection and provides a resource of small molecules to study ZIKV pathogenesis.

To understand how nuclear processes involving DNA are regulated, knowledge of the determinants of chromatin condensation is required. From recent structural studies it has been concluded that the formation of the 30-nm chromatin fiber does not require the linker histone. Here, by comparing the linker histone-dependent compaction of long, reconstituted nucleosome arrays with different nucleosome repeat lengths (NRLs), 167 and 197 bp, we establish that the compaction behavior is both NRL-and linker histone-dependent. Only the 197-bp NRL array can form 30-nm higher-order chromatin structure. Importantly for understanding the regulation of compaction, this array shows a cooperative linker histone-dependent compaction. The 167-bp NRL array displays a limited linker histone-dependent compaction, resulting in a thinner and topologically different fiber. These observations provide an explanation for the distribution of NRLs found in nature.30-nm fiber ͉ electron microscopy ͉ heterochromatin ͉ nucleosome array reconstitution ͉ sedimentation velocity analysis D uring the past decade it has emerged that the packaging of eukaryotic DNA by histones into chromatin is a key regulator of nuclear processes involving DNA, such as transcription and replication. Although the structure of the first level of DNA folding, the nucleosome core, is known at atomic resolution (1, 2), the structure of the second level of folding, whereby a string of nucleosomes folds into a fiber with an approximate diameter of 30 nm (the 30-nm chromatin fiber) remains undetermined (3). Early evidence for the presence of a 30-nm chromatin fiber in vivo came from EM analysis of Balbiani ring genes in Chironomus tentans (4) and x-ray diffraction studies of nuclei that show spacings of 30-40 nm (5).The structure of the 30-nm chromatin fiber is controversial (reviewed in ref.3). Recent structural analyses using in vitroreconstituted model nucleosome arrays based on the strong 601 DNA nucleosome positioning sequence (6) have led to the proposal of two models for the 30-nm chromatin fiber that differ in topology, dimension, and nucleosome packing density. The first model was constructed by using the crystal structure at 9 Å of a tetra-nucleosome core array and is of a two-start helix type (7). It is based on a zigzag arrangement of nucleosome cores that stack on top of each other and has a 24-to 25-nm diameter with a packing density of five to six nucleosomes per 11 nm. The second model was derived from tight constraints obtained from measurements of the physical dimensions of long nucleosome arrays visualized by EM (8). It is of the one-start helix type in which nucleosomes from adjacent gyres are interdigitated. It has a diameter of 34 nm with a packing density of 11 nucleosomes per 11 nm.The key difference between the two 30-nm chromatin fiber models is that the interdigitated model was derived from nucleosome arrays saturated with linker histone, whereas the two-start helix model was derived from a tetra-nucleosome core array in the absence of linker histone...