Multi-omics analysis of chromatin accessibility and interactions with transcriptome by HiCAR

Abstract

High-order chromatin organization plays a central role in regulating spatial-temporal gene expression by facilitating or constraining the interactions between cis-regulatory elements (cREs). cREs are often the accessible DNA sequences and can be identified at genome-wide scale with assays such as ATAC-seq, DHS-seq, and FAIRE-seq. However, it remains technically challenging to comprehensively identify the long-range interactions that occur between cREs, especially using low-input cells and in a cost effective manner. Here, we report HiCAR, Hi gh-throughput C hromosome conformation capture on A ccessible DNA with m R NA-seq co-assay, which enables simultaneous mapping of chromatin accessibility and cRE anchored chromatin contacts. Notably, using the same input material, HiCAR also yields high-quality transcriptome data that represents the functional outputs of chromatin accessibility and interaction. Unlike immunoprecipitation-based methods such as HiChIP, PLAC-seq, and ChIA-PET, HiCAR does not require target-specific antibodies and thus can capture cis-regulatory contacts anchored on the accessible DNA regions associated with multiple epigenetic modifications and transcription factor binding. We compared HiCAR to another technology designed to capture interactions between accessible chromatin regions, called Trac-looping, and found that HiCAR yielded much more informative long-range cis- reads at similar sequencing depth, and requires far fewer cells as input. We applied HiCAR to H1 human embryonic stem cells (hESC) and identified 46,792 open-chromatin anchored loops at 5Kb resolution. Interestingly, we found that the poised cREs form extensive and significant chromatin interactions comparable to the active cREs. We further showed that the spatial interactive activity of cREs do not correlate with their transcriptional activity, enhancer activity, and chromatin accessibility. Additionally, we identified 2,096 super interactive regulatory (SINTER) loci showing abnormally high levels of chromatin interactivity and associated with unique epigenetic features. In summary, HiCAR is a robust, sensitive, and cost effective multi-omics coassay that can be used to study chromatin structure and function as well as gene expression.

Spatial chromatin accessibility sequencing resolves high-order spatial interactions of epigenomic markers

“…Recently, there has been an increasing interest in applying methylation labeling and nanopore sequencing for the analysis of chromatin accessibility at a single-molecule level [10][11][12][13][14]24 . In this Compared with the output of the comparable techniques Trac-loop 21 , Hi-CAR 23 , and NicE-C 25 , which also capture the accessible chromatin conformation, the SCA-seq preserved more multi-omics information, for example, CpG methylation epigenetic marks, chromatin inaccessibility, and high-order chromatin conformation data (Fig 1b).…”

“…However, chromatin has a higher-order organization, and the linearized two-dimensional map of chromatin accessibility does not fully reflect the reality. Some advanced approaches, such as Trac-looping 11 , OCEAN-C 12 , and HiCAR 13 , could decrease the need for Hi-C data 14 by enrichment of open chromatin regions by combining transposons and proximity ligation. However, the losses at the single-molecule resolution level and the condensed domains restrict the possibility to observe dynamic changes in chromatin structure in three-dimensional space.…”

“…Most of these techniques do not allow obtaining epigenome information in the process or examining chromatin conformation with multi-omics. Some advanced approaches, such as Trac-looping 21 , OCEAN-C 22 , and HiCAR 23 , can capture open chromatin and chromatin conformation information simultaneously by enrichment of accessible chromatin regions through solubility or transposons. These advanced approaches were originally developed to enrich the accessible chromatin and efficiently observe the interactions of cis-regulatory elements (cREs).…”

Spatial chromatin accessibility sequencing resolves next-generation genome architecture

“…1a,b). Compared with the competing techniques Trac-loop 11 , Hi-CAR 13 and NicE-C 15 , which also captured the accessible chromatin conformation, the SCA-seq could reserve more multi-omics information, for example, CpG methylation epigenetic marks, chromatin inaccessibility, and high-order chromatin conformation (Fig. 1b).…”

“…However, chromatin has a higher-order organization, and the linearized two-dimensional map of chromatin accessibility does not fully re ect reality. Some advanced approaches, such as Trac-looping 11 , OCEAN-C 12 , and HiCAR 13 , could capture open chromatin and Hi-C 14 information simultaneously by enrichment of accessible chromatin regions through the combination of transposons and proximity ligation. However, the losses at the singlemolecule resolution level and the condensed chromatin regions restrict the possibility to observe dynamic changes in chromatin structure in three-dimensional space.…”