Conserved facultative heterochromatin across cell types identify regulatory sequences underpinning cell identity and disease

Cellular diversity is a product of evolution acting to establish divergent regulatory programs from a common genome. Here, we use cross-cell-type epigenetic conservation to gain insight into the impact of selective constraints on genome function and phenotypic variation. By comparing chromatin accessibility across hundreds of diverse cell-types, we identify 1.4% of the human genome safeguarded by conserved domains of facultative heterochromatin, which we term regions under “cellular constraint”. We calculate single-base resolution cellular constraint scores and demonstrate robust prediction of functionally important coding and non-coding loci in a cell-type-, trait-, and disease-agnostic manner. Cellular constraint annotation enhances causal variant identification, drug discovery, and clinical diagnostic predictions. Furthermore, cell-constrained sequences share signals of positive and negative selection, suggesting a dynamic role in influencing human fitness and adaptation. Overall, this study demonstrates that evolutionary chromatin dynamics can be leveraged to inform the translation of genetic discoveries into effective biological, therapeutic, and clinical outcomes. Overall design: Hashtag barcoded WTC CRISPRi (Control) and RP11-175E9.1-g1 CRISPRi WTC (NKX2-6-AS1 K/D) hiPSCs were differentiated and on day 6 were pooled together with other samples for sequencing. Single live cells were collected and FACs sorted for a multiplexed scRNA-seq run.