Cohesin complexes mediate sister-chromatid cohesion in dividing cells but may also contribute to gene regulation in postmitotic cells. How cohesin regulates gene expression is not known. Here we describe cohesin-binding sites in the human genome and show that most of these are associated with the CCCTC-binding factor (CTCF), a zinc-finger protein required for transcriptional insulation. CTCF is dispensable for cohesin loading onto DNA, but is needed to enrich cohesin at specific binding sites. Cohesin enables CTCF to insulate promoters from distant enhancers and controls transcription at the H19/IGF2 (insulin-like growth factor 2) locus. This role of cohesin seems to be independent of its role in cohesion. We propose that cohesin functions as a transcriptional insulator, and speculate that subtle deficiencies in this function contribute to 'cohesinopathies' such as Cornelia de Lange syndrome.

We describe a simple illumination method of fluorescence microscopy for molecular imaging. Illumination by a highly inclined and thin beam increases image intensity and decreases background intensity, yielding a signal/background ratio about eightfold greater than that of epi-illumination. A high ratio yielded clear single-molecule images and three-dimensional images using cultured mammalian cells, enabling one to visualize and quantify molecular dynamics, interactions and kinetics in cells for molecular systems biology.

Using deep sequencing (deepCAGE), the FANTOM4 study measured the genome-wide dynamics of transcription-start-site usage in the human monocytic cell line THP-1 throughout a time course of growth arrest and differentiation. Modeling the expression dynamics in terms of predicted cis-regulatory sites, we identified the key transcription regulators, their time-dependent activities and target genes. Systematic siRNA knockdown of 52 transcription factors confirmed the roles of individual factors in the regulatory network. Our results indicate that cellular states are constrained by complex networks involving both positive and negative regulatory interactions among substantial numbers of transcription factors and that no single transcription factor is both necessary and sufficient to drive the differentiation process.

In response to interferon-γ (IFN-γ), Stat1 is tyrosine phosphorylated and translocates to the nucleus where it activates transcription. In this study, we identified factors which mediate the nuclear import of Stat1. Tyrosine-phosphorylated Stat1 associated with the β subunit (a 97 kDa component) of the nuclear poretargeting complex via the NPI-1 family, but not the Rch1 family, of α subunit (a 58 kDa component) as a result of IFN-γ stimulation. Antibodies against NPI-1 or β subunit consistently inhibited the IFN-γ-dependent nuclear import of Stat1 in living cells, although antibodies reactive to Rch1 had no effect. Solution binding assays with deletion mutants of NPI-1 showed that the Stat1-binding domain of NPI-1 was located in the carboxy-terminal region, which is clearly distinct from the SV40 large T antigen nuclear localization signal (NLS)-binding region. These results indicate that the extracellular signal-dependent nuclear transport of Stat1 is mediated by NPI-1, but not Rch1, in conjunction with β subunit, and that these factors participate in, not only constitutive, but also the conditional nuclear import of proteins.