Summary
Spatial genome organization and its effect on transcription remains a fundamental question. We applied an advanced ChIA-PET strategy to comprehensively map higher-order chromosome folding and specific chromatin interactions mediated by CTCF and RNAPII with haplotype specificity and nucleotide resolution in different human cell lineages. We find that CTCF/cohesin-mediated interaction anchors serve as structural foci for spatial organization of constitutive genes concordant with CTCF-motif orientation, whereas RNAPII interacts within these structures by selectively drawing cell-type-specific genes towards CTCF-foci for coordinated transcription. Furthermore, we show that haplotype-variants and allelic-interactions have differential effects on chromosome configuration influencing gene expression and may provide mechanistic insights into functions associated with disease susceptibility. 3D-genome simulation suggests a model of chromatin folding around chromosomal axes, where CTCF is involved in defining the interface between condensed and open compartments for structural regulation. Our 3D-genome strategy thus provides unique insights in the topological mechanism of human variations and diseases.
The acquisition of proper dendrite morphology is a crucial aspect of neuronal development towards the formation of a functional network. The role of the extracellular matrix and its cellular receptors in this process has remained enigmatic. We report that the CD44 adhesion molecule, the main hyaluronan receptor, is localized in dendrites and plays a crucial inhibitory role in dendritic tree arborization in vitro and in vivo. This novel function is exerted by the activation of Src tyrosine kinase, leading to the alteration of Golgi morphology. The mechanism operates during normal brain development, but its inhibition might have a protective influence on dendritic trees under toxic conditions, during which the silencing of CD44 expression prevents dendritic shortening induced by glutamate exposure. Overall, our results indicate a novel role for CD44 as an essential regulator of dendritic arbor complexity in both health and disease.
The human genome is extensively folded into 3-dimensional organization. However, the detailed 3D chromatin folding structures have not been fully visualized due to the lack of robust and ultra-resolution imaging capability. Here, we report the development of an electron microscopy method that combines serial block-face scanning electron microscopy with in situ hybridization (3D-EMISH) to visualize 3D chromatin folding at targeted genomic regions with ultra-resolution (5 × 5 × 30 nm in xyz dimensions) that is superior to the current super-resolution by fluorescence light microscopy. We apply 3D-EMISH to human lymphoblastoid cells at a 1.7 Mb segment of the genome and visualize a large number of distinctive 3D chromatin folding structures in ultra-resolution. We further quantitatively characterize the reconstituted chromatin folding structures by identifying sub-domains, and uncover a high level heterogeneity of chromatin folding ultrastructures in individual nuclei, suggestive of extensive dynamic fluidity in 3D chromatin states.
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