Histone H3 trimethylation of lysine 9 (H3K9me3) and proteins of the heterochromatin protein 1 (HP1) family are hallmarks of heterochromatin, a state of compacted DNA essential for genome stability and long-term transcriptional silencing. The mechanisms by which H3K9me3 and HP1 contribute to chromatin condensation have been speculative and controversial. Here we demonstrate that human HP1β is a prototypic HP1 protein exemplifying most basal chromatin binding and effects. These are caused by dimeric and dynamic interaction with highly enriched H3K9me3 and are modulated by various electrostatic interfaces. HP1β bridges condensed chromatin, which we postulate stabilizes the compacted state. In agreement, HP1β genome-wide localization follows H3K9me3-enrichment and artificial bridging of chromatin fibres is sufficient for maintaining cellular heterochromatic conformation. Overall, our findings define a fundamental mechanism for chromatin higher order structural changes caused by HP1 proteins, which might contribute to the plastic nature of condensed chromatin.

SNARE proteins mediate membrane fusion in eukaryotic cells. They contain conserved SNARE motifs that are usually located adjacent to a C-terminal transmembrane domain. SNARE motifs spontaneously assemble into four helix bundles, with each helix belonging to a different subfamily. Liposomes containing SNAREs spontaneously fuse with each other, but it is debated how the SNAREs are distributed between the membranes. Here, we report that the SNAREs mediating homotypic fusion of early endosomes fuse liposomes in five out of seven possible combinations, in contrast to previously studied SNAREs involved in heterotypic fusion events. The crystal structure of the early endosomal SNARE complex resembles that of the neuronal and late endosomal complexes, but differs in surface side-chain interactions. We conclude that homotypic fusion reactions may proceed with multiple SNARE topologies, suggesting that the conserved SNARE structure allows for flexibility in the initial interactions needed for fusion.

We present results of a two-photon fluorescence-correlation study carried out with glycosylated and untreated 20 nm fluorescing spheres that interacted with the carbohydrate-binding proteins soybean agglutinin (SBA) and concanavalin A (Con A). The assay principle allows protein-carbohydrate binding interactions to be determined without protein labeling. This assay might serve as a simple model system for studying physical and chemical interactions between proteins and carbohydrates, for example, at cell or virus surfaces. In experiments with galactosylated 20 nm beads and SBA, several stages of protein-carbohydrate interactions could be clearly distinguished. Initially, only a few lectins bound to the nanospheres. At higher lectin concentrations polymerization occurred, and aggregates consisting of about 2.6 x 10(5) glycosylated nanospheres were formed. At very high lectin concentrations, the degree of polymerization dropped, and the size of single SBA-covered nanospheres increased to approximately 40 nm. When Con A was used instead of SBA, a significantly smaller degree of aggregation (4 x 10(4) spheres) was obtained. Treatment of unglycosylated 20 nm beads with SBA as a negative control sample resulted in a much lower unspecific aggregation (5 x 10(3) spheres). The assay principle can thus help to elucidate relative binding affinities.

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Scanning electron microscope images show that it is easy to generate nanopores on polycarbonate membranes with well-defined pore diameters by ion-track perforation and subsequent magnetron sputtering with metal. The size reduction of the nanopores during sputtering with gold is a linear function of time. Images of different angles and from the bottom side of the membrane show that the channels are the smallest very close to the surface of the metal layer, have a conelike shape, and reach about half as much into the polymer membranes as the metal-layer thickness. This topographical pore shape is ideal for use as optically coherent near-field sources in deep-nulling microscopy. We present the first results of significantly improved nulling stabilization in the presence (<2 nm optical pathway difference) and the absence (<0.6 nm optical pathway difference) of the nanoapertures in the focal region of a deep-nulling microscope.

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