Chromatin structure plays a fundamental role in the regulation of nuclear processes such as DNA transcription, replication, recombination, and repair. Despite considerable efforts during three decades, the structure of the 30-nm chromatin fiber remains controversial. To define fiber dimensions accurately, we have produced very long and regularly folded 30-nm fibers from in vitro reconstituted nucleosome arrays containing the linker histone and with increasing nucleosome repeat lengths (10 to 70 bp of linker DNA). EM measurements show that the dimensions of these fully folded fibers do not increase linearly with increasing linker length, a finding that is inconsistent with two-start helix models. Instead, we find that there are two distinct classes of fiber structure, both with unexpectedly high nucleosome density: arrays with 10 to 40 bp of linker DNA all produce fibers with a diameter of 33 nm and 11 nucleosomes per 11 nm, whereas arrays with 50 to 70 bp of linker DNA all produce 44-nm-wide fibers with 15 nucleosomes per 11 nm. Using the physical constraints imposed by these measurements, we have built a model in which tight nucleosome packing is achieved through the interdigitation of nucleosomes from adjacent helical gyres. Importantly, the model closely matches raw image projections of folded chromatin arrays recorded in the solution state by using electron cryo-microscopy.chromatin structure ͉ electron microscopy ͉ linker histone ͉ reconstitution E ukaryotic chromosomes have a compact structure in which linear nucleosome arrays are first folded into a fiber of around 30-nm diameter (1, 2). The fundamental repeating unit of chromatin, the nucleosome core particle, organizes 147 bp of DNA in 1.7 left-handed superhelical turns around an octamer of the four core histones (H2A, H2B, H3, and H4) (3-5). Linker histone (H1͞H5) binding organizes an additional 20 bp of DNA to complete the nucleosome containing 167 bp of DNA (6, 7). Such binding determines the geometry of the DNA entering and exiting the nucleosome core particle (8). In nucleosome arrays, adjacent nucleosomes are separated by linker DNA, varying in length between 0 and 80 bp in a tissue-and species-specific manner (9, 10). In vitro, linear nucleosome arrays fold into the ''30-nm'' fiber upon increasing ionic strength (11) in a process that depends on both the integrity of the core histone N-terminal tails (12, 13) and the presence of the linker histone (14,15).During the past three decades evidence from EM (14-23), x-ray and neutron scattering (24-27), electric and photochemical dichroism (28-31), sedimentation analysis (32-35), nuclease digestion (6, 9, 36), and x-ray crystallography (4, 5, 37, 38) has led to the proposal of a number of different models for the 30-nm fiber. These models fall into two main classes: the one-start helix or solenoid models, and the two-start helix models. The solenoid models are comprised of simple one-start helices in which successive nucleosomes are adjacent in the filament and connected by linker DNA that bends into t...

SummaryHistone deacetylase enzymes (HDACs) are emerging cancer drug targets. They regulate gene expression by removing acetyl groups from lysine residues in histone tails resulting in chromatin condensation. The enzymatic activity of most class I HDACs requires recruitment to corepressor complexes. We report the first structure of an HDAC:corepressor complex - HDAC3 with the deacetylase-activation-domain (DAD) from the SMRT corepressor. The structure reveals two remarkable features. First the SMRT-DAD undergoes a large structural rearrangement on forming the complex. Second there is an essential inositol tetraphosphate molecule, Ins(1,4,5,6)P4, acting as an ‘intermolecular glue’ between the two proteins. Assembly of the complex is clearly dependent on the Ins(1,4,5,6)P4, which may act as a regulator – potentially explaining why inositol phosphates and their kinases have been found to act as transcriptional regulators. This mechanism for the activation of HDAC3 appears to be conserved in class I HDACs from yeast to man and opens novel therapeutic opportunities.

SummaryClass I histone deacetylases (HDAC1, HDAC2, and HDAC3) are recruited by cognate corepressor proteins into specific transcriptional repression complexes that target HDAC activity to chromatin resulting in chromatin condensation and transcriptional silencing. We previously reported the structure of HDAC3 in complex with the SMRT corepressor. This structure revealed the presence of inositol-tetraphosphate [Ins(1,4,5,6)P4] at the interface of the two proteins. It was previously unclear whether the role of Ins(1,4,5,6)P4 is to act as a structural cofactor or a regulator of HDAC3 activity. Here we report the structure of HDAC1 in complex with MTA1 from the NuRD complex. The ELM2-SANT domains from MTA1 wrap completely around HDAC1 occupying both sides of the active site such that the adjacent BAH domain is ideally positioned to recruit nucleosomes to the active site of the enzyme. Functional assays of both the HDAC1 and HDAC3 complexes reveal that Ins(1,4,5,6)P4 is a bona fide conserved regulator of class I HDAC complexes.

The nuclear receptor peroxisome proliferator-activated receptor-gamma (PPARgamma) has important roles in adipogenesis and immune response as well as roles in both lipid and carbohydrate metabolism. Although synthetic agonists for PPARgamma are widely used as insulin sensitizers, the identity of the natural ligand(s) for PPARgamma is still not clear. Suggested natural ligands include 15-deoxy-delta12,14-prostaglandin J2 and oxidized fatty acids such as 9-HODE and 13-HODE. Crystal structures of PPARgamma have revealed the mode of recognition for synthetic compounds. Here we report structures of PPARgamma bound to oxidized fatty acids that are likely to be natural ligands for this receptor. These structures reveal that the receptor can (i) simultaneously bind two fatty acids and (ii) couple covalently with conjugated oxo fatty acids. Thermal stability and gene expression analyses suggest that such covalent ligands are particularly effective activators of PPARgamma and thus may serve as potent and biologically relevant ligands.