Tracing the evolution of single-cell 3D genomes in Kras-driven cancers

Although three-dimensional (3D) genome structures are altered in cancer cells, little is known about how these changes evolve and diversify during cancer progression. Leveraging genome-wide chromatin tracing to visualize 3D genome folding directly in tissues, we generated 3D genome cancer atlases of oncogenic Kras-driven murine lung and pancreatic adenocarcinoma. Our data reveal stereotypical, non-monotonic, and stage-specific alterations in 3D genome folding compaction, heterogeneity, and compartmentalization as cancers progress from normal to preinvasive and ultimately to invasive tumors, discovering a potential structural bottleneck in early tumor progression. Remarkably, 3D genome architectures distinguish morphologic cancer states in single cells, despite considerable cell-to-cell heterogeneity. Gene-level analyses of evolutionary changes in 3D genome compartmentalization not only showed that compartment-associated genes are more homogeneously regulated but also elucidated prognostic and dependency genes in lung adenocarcinoma and a previously unappreciated role for polycomb-group protein Rnf2 in 3D genome regulation. Our results demonstrate the utility of mapping the single-cell cancer 3D genome in tissues and illuminate its potential to identify new diagnostic, prognostic, and therapeutic biomarkers in cancer. CUT&RUN of RNF2, H3K4me3, H3K27me3, H2AK119ub, BMI-1, RNA Pol II p-ser5 and IgG in the mouse KP LUAD cell line (31671) stably transfected with shRNAs targeting Rnf2 (shRnf2-3) and a non-targeting control (shNTC). The cell line was derived from an autochthonous KrasLSL-G12D/WT; Trp53flox/flox mouse administered with Adeno-Cre.