HiCuT: An efficient and low input method to identify protein-directed chromatin interactions

3D genome organization regulates gene expression, and disruption of these long-range (>20kB) DNA-protein interactions results in pathogenic phenotypes. Chromosome conformation methods in conjunction with chromatin immunoprecipitation were used to decipher protein-directed chromatin interactions. However, these methods required abundant starting material (>500,000 cells), sizable number of sequencing reads (>100 million reads), and elaborate data processing methods to reduce background noise, which limited their use in primary cells. Hi-C Coupled chromatin cleavage and Tagmentation (HiCuT) is a new transposase-assisted tagmentation method that generates high-resolution protein directed long-range chromatin interactions as efficiently as existing methods, HiChIP and ChIA-PET, despite using 100,000 cells (5-fold less) and 12 million sequencing reads (8-fold fewer). Moreover, HiCuT generates high resolution fragment libraries with low background signal that are easily interpreted with minimal computational processing. We used HiCuT in human primary skin cells to link previously identified single nucleotide polymorphisms (SNPs) in skin disease to candidate genes and to identify functionally relevant transcription factors in an unbiased manner. HiCuT broadens the capacity for genomic profiling in systems previously unmeasurable, including primary cells, human tissue samples, and rare cell populations, and may be a useful tool for all investigators studying human genetics and personalized epigenomics. Author summaryDNA is precisely packaged and organized within a nucleus to regulate gene expression. Altering this structure results in disease and developmental abnormalities. Current methods to probe 3D genome organization require a minimum of 500,000 cells, 100 million sequencing reads, and elaborate computational genomics skills. This limits general adoption and prevents use in small populations of cells, including primary tissues. We report a new method called Hi-C Coupled chromatin cleavage and Tagmentation (HiCuT). This method couples current 3D genome methods in conjunction with target specific tagmentation, an enzyme-assisted approach to cut and tag DNA. We benchmarked HiCuT against existing methods and found similar efficiency and specificity in assessing target specific 3D genome organization despite reducing assay requirements to 100,000 cells (5-fold less) and 12 million sequencing reads (8-fold fewer). Strikingly, HiCuT data exhibited reduced background noise and required minimal computational processing. Taken together, HiCuT broadens the capacity for 3D genome profiling in cell populations previously unmeasurable, including primary cells and human tissues, reduces sequencing costs, and lowers the need for computational expertise. HiCuT will benefit all investigators studying gene regulation and disease pathophysiology.