Cut-C: cleavage under tethered nuclease for conformational capture

Abstract

Background Deciphering the 3D structure of the genome is essential for elucidating the regulatory mechanisms of gene expression in detail. Existing methods, such as chromosome conformation capture (3C) and Hi-C have enabled the identification of novel aspects of chromatin structure. Further identification of protein-centric chromatin conformation is enabled by coupling the Hi-C procedure with a conventional chromatin immunoprecipitation assay. However, these methods are time-consuming and require independent methods for validation. Results To simultaneously identify protein-centric chromatin conformation and target protein localization, we have developed Cut-C, a method that combines antibody-mediated cleavage by tethered nuclease with chromosome conformation capture to identify chromatin interactions mediated by a protein of interest. Applying Cut-C to H3K4me3, a histone modification enriched at active gene promoters, we have successfully identified chromatin loops mediated by H3K4me3 along with the genome-wide distribution of H3K4me3. Cut-C also identified chromatin loops mediated by CTCF, validating the general applicability of the method. Conclusions Cut-C identifies protein-centric chromatin conformations along with the genome-wide distribution of target proteins using simple procedures. The simplified protocol will improve the efficiency of analysing chromatin conformation using precious materials, such as clinical samples. Electronic supplementary material The online version of this article (10.1186/s12864-019-5989-2) contains supplementary material, which is available to authorized users.

ZNF143 in Chromatin Looping and Gene Regulation

“…Skene et al have showed that their new in situ methods, such as cleavage under targets and release using nuclease (CUT&RUN) and cleavage under targets and tagmentation (CUT&Tag), will be viewed as a cost-effective and versatile alternative to ChIP because of low backgrounds, which requiring only ∼1/10th the sequencing depth as ChIP (Skene and Henikoff, 2017;Skene et al, 2018;Kaya-Okur et al, 2019;Meers et al, 2019). Based on these methods, Shimbo et al (2019) have developed cleavage under tethered nuclease for conformational capture (Cut-C) technology to identify chromatin interactions mediated by a protein of interest along with the genome-wide distribution of the target proteins. Thus, using these latest technologies, we may be clearly captured the accuracy of chromatin loops mediated by ZNF143 in a genome-wide scale.…”

Plant 3D genomics: the exploration and application of chromatin organization

“…This article is protected by copyright. All rights reserved combines antibody-mediated cleavage by tethered nuclease with chromosome conformation capture (Shimbo et al, 2019); Capture Hi-C: a method that combines Hi-C with the hybridization-based capture of targeted genomic regions (Mifsud et al, 2015); In situ Hi-C: a method in which DNA-DNA proximity ligation is performed in intact nuclei (Rao et al, 2014); Micro-C: a method in which chromatin is fragmented into mononucleosomes using micrococcal nuclease (Hsieh et al, 2015); Dnase Hi-C: a method in which chromatin is fragmented by DNase I (Ma et al, 2015); DLO Hi-C: Digestion-ligation-only Hi-C technology that requires two rounds of digestion and ligation, without the need for biotin labeling and pulldown (Lin et al, 2018); BAT Hi-C: Bridge linker-Alul-Tn5 Hi-C method that combines Alul fragmentation with biotinylated linker-mediated proximity ligation ; BL-Hi-C: Bridge Linker-Hi-C that requires restriction enzyme targeting and two-step proximity ligation (Liang et al, 2017); Trac-looping: transposase-mediated analysis of chromatin looping (Lai et al, 2018); Methyl-HiC: a method that combines Hi-C and DNA methylation detection technology (Li et al, 2019b); OCEAN-C: open chromatin enrichment and network Hi-C ; Single cell Hi-C: a method to perform Hi-C in an individual nucleus (Nagano et al, 2013). Asterisks show the methods that have been applied to plants.…”

Understanding 3D Genome Organization and Its Effect on Transcriptional Gene Regulation Under Environmental Stress in Plant: A Chromatin Perspective

“…The recently developed techniques like Trac-looping (transposase-mediated analysis of chromatin looping) for simultaneous detection of multiscale genome-wide chromatin interactions among regulatory elements and chromatin accessibility ( Lai et al, 2018 ), and OCEAN Hi-C (open chromatin enrichment and network Hi-C) for antibody-independent mapping of global open chromatin interactions ( Li T. et al, 2018 ) were used to decipher the chromatin architecture. More recently, Cut-C combined antibody-mediated cleavage by tethered nuclease with chromosome conformation capture to identify chromatin interactions mediated by a protein of interest ( Shimbo et al, 2019 ). Cut-C identifies protein-centric chromatin conformations along with the genome-wide distribution of target proteins using a simple procedure.…”

“…Cut-C identifies protein-centric chromatin conformations along with the genome-wide distribution of target proteins using a simple procedure. Applying Cut-C to a histone modification (H3K4me3) enriched at active gene promoters, Shimbo et al (2019) could successfully identify the chromatin loops mediated by H3K4me3 along with the genome-wide distribution of H3K4me3. Further, methyl Hi-C (DNA methylation detection combined with Hi-C) for simultaneous capture of chromosome conformation and DNA methylome was used to delineate the DNA methylation profile and chromatin architecture of a cell ( Li G. et al, 2019 ).…”

“…More recently, Cut-C combined antibody-mediated cleavage by tethered nuclease with chromosome conformation capture to identify chromatin interactions mediated by a protein of interest (Shimbo et al, 2019). Cut-C identifies protein-centric chromatin conformations along with the genome-wide distribution of target proteins using a simple procedure.…”