Hi-C-based assembly of a highly rearranged human cell line model system. (A) Schematic overview of Complex Alterations after Selection and Transformation (CAST) coupled with in situ Hi-C. TP53-deficient RPE-1 cells are perturbed with doxorubicin (DOX), followed by single-cell sorting and expansion. Surviving populations are screened for genomic rearrangements with low-depth WGS. Hi-C libraries from control (maternal) and treated (daughter) clones are sequenced to identify acquired and drug-induced rearrangements. (B) Annotation of fragment size and edge positions. The latter is annotated at peaks of ectopic contacts between two distal sites. Duplicated regions, highlighted at fragments F and G, exhibit distinct interaction patterns and produce nonorthogonal ectopic interactions, as revealed by the interaction profile between fragments C-F and B-G, respectively. The genomic regions depicted on the Hi-C maps correspond to the highlighted regions on the chromosome ideograms. (C, upper) Following the above-described annotation strategy, we create a graph of nodes (fragments) and edges (rearrangements) for the ectopic interactions of panel B. (Lower) Traversing the graph produces the derivative assembly. (−1) denotes fragment inversion. Duplicated regions that are shared between fragments A-B and F-G are colored in pink. (D) The Hi-C map depicts rearrangements within and across Chromosomes 2, 12, and 22. Straight diagonal lines on the Hi-C map trace the position of the SV calls back to the reference genome. Following the graph method in B–D, we annotated 12 rearranged DNA fragments (A–L) and their juxtapositions, represented as green and yellow rectangles and directed arches, respectively. The telomere of Chromosome 2q is connected to 12 and 22, observed as stripes protruding from the 2q telomere to the q-arm of 12 and 22. (E) Using Chromosome 2 as a starting point, we walk along the annotated edges and segments to produce the assembly of the derivative 2-12-22 chromosome.
