Reconstruction of diploid higher-order human 3D genome interactions from noisy Pore-C data using Dip3D.

Differential high-order chromatin interactions between homologous chromosomes affect many biological processes. Traditional chromatin conformation capture genome analysis methods mainly identify two-way interactions and cannot provide comprehensive haplotype information, especially for low-heterozygosity organisms such as human. Here, we present a pipeline of methods to delineate diploid high-order chromatin interactions from noisy Pore-C outputs. We trained a previously published single-nucleotide variant (SNV)-calling deep learning model, Clair3, on Pore-C data to achieve superior SNV calling, applied a filtering strategy to tag reads for haplotypes and established a haplotype imputation strategy for high-order concatemers. Learning the haplotype characteristics of high-order concatemers from high-heterozygosity mouse allowed us to devise a progressive haplotype imputation strategy, which improved the haplotype-informative Pore-C contact rate 14.1-fold to 76% in the HG001 cell line. Overall, the diploid three-dimensional (3D) genome interactions we derived using Dip3D surpassed conventional methods in noise reduction and contact distribution uniformity, with better haplotype-informative contact density and genomic coverage rates. Dip3D identified previously unresolved haplotype high-order interactions, in addition to an understanding of their relationship with allele-specific expression, such as in X-chromosome inactivation. These results lead us to conclude that Dip3D is a robust pipeline for the high-quality reconstruction of diploid high-order 3D genome interactions.