Andrew B. Stergachis

Enhancer Tuning by Sequence-Specific Repressors

Enhancers integrate combinatorial inputs from sequence-specific transcription factors (TFs) and their activity must be calibrated to achieve precise spatiotemporal control of transcript dosage. Here we demonstrate that the sequence-specific repressors SNAI1 and SNAI2 (i.e. SNAIL and SLUG) quantitatively tune enhancer activity. In human neural crest cells, SNAI1/2 occupy a subset of active enhancers, where their depletion increases H3K27ac, chromatin accessibility, and enhancer regulatory potential. Changes in SNAI1/2 binding motifs contribute to enhancer divergence between human and chimpanzee, suggesting an evolutionary role for the repressor-mediated tuning. Single-molecule chromatin profiling using Deaminase-Assisted Fiber-seq (DAF-seq) reveals that individual enhancers toggle between an ensemble of open and nucleosome-dense chromatin states. While transcriptional activators increase the fraction of the open states, SNAI1/2 shift the equilibrium toward nucleosome-occupied states. This impedes binding of activator TFs, without fully repressing the enhancer. We propose that SNAI1/2 function as a molecular dimmer switch—modulating nucleosome dynamics to calibrate enhancer output.

CHROMATIN SEQUENCING Single-molecule regulatory architectures captured by chromatin fiber sequencing

Inverted logic of ecDNA localization converts nuclear periphery into oncogenic transcription hubs in cancer

Extrachromosomal DNA (ecDNA) is a major driver of cancer pathogenesis, however, its spatial organization in the nucleus is not currently understood. Here we show that ecDNAs, in contrast to chromosomal DNA, preferentially localize to the nuclear periphery yet resist peripheral gene silencing. Single-molecule multi-omic sequencing and imaging-based CRISPR screening demonstrate enhanced ecDNA hub formation, H3K27ac and H3K9me3 bivalency, late DNA replication, long-range CpG hypomethylation, and increased transcriptional output when ecDNAs localize to the nuclear periphery. EcDNA anchors to the nuclear lamina through H3K9me3-dependent interactions with the Lamin B Receptor (LBR), which if interrupted, results in inward migration of ecDNA particles, reduced ecDNA congregation, and decreased transcription of ecDNA-encoded genes. These results reveal inherent advantages of the spatial organization of ecDNA that are not available to chromosomal DNA, enhancing ecDNA congregation and amplifying oncogenic transcriptional output, by localizing to the nuclear periphery of cancer cells.

Integrated map of somatic mosaicism across human tissues in 25 individuals

Although all cells in the body descend from one genome, they accumulate distinct genetic and epigenetic changes over a lifetime, producing a mosaic of somatic variation that can shape development, aging, and disease. This mosaicism is often studied in isolation, leaving unclear how these forms relate within and between individuals. Here, we present the first integrated analysis of the Somatic Mosaicism across Human Tissues (SMaHT) Network’s production resource, profiling up to 20 tissues from 25 donors using short- and long-read, duplex, single-cell, transcriptomic, and epigenomic sequencing, alongside donor-specific near-telomere-to-telomere assemblies. Somatic mutation burden cannot be captured by a single data type or metric, as tissues accumulate distinct variant classes largely independently of one another. Long-read and single-cell data resolved cell-type-specific mutational processes, traced mobile element insertions to source loci, and revealed the developmental timing and functional consequences of individual mutations. Donor-specific assemblies uncovered elevated mutation rates within centromeres and segmental duplications inaccessible to standard reference genomes, while haplotype-resolved chromatin and methylation data showed that nongenetically-deterministic epigenetic states are pervasive across tissues. Together, these findings provide an integrated, multi-scale portrait of somatic mosaicism across the human body, establishing a baseline against which its contributions to aging and disease can be measured.

A complete diploid human genome benchmark for personalized genomics

SUMMARY Human genome sequencing typically relies on mapping reads to a reference genome to call variants, but this approach introduces technical biases, excluding duplicated and structurally polymorphic regions of the genome. To overcome this, we present a telomere-to-telomere genome benchmark with near-perfect accuracy across 99.4% of the diploid HG002 genome. This benchmark adds 701.4 Mb of autosomal sequence and both sex chromosomes (216.8 Mb), which were absent from prior benchmarks. We annotated genes and repeats on both haplotypes, including 19,956 protein-coding genes on the maternal haplotype and 19,190 on the paternal haplotype, and developed new methods to measure the accuracy of reads, phased variant call sets, and assemblies against a diploid reference. Genome-wide analyses show that de novo assembly resolves 2%–7% more sequence and outperforms variant calling accuracy by an order of magnitude, expanding the reach of genomic medicine to the entire genome and enabling a new era of personalized genomics.

Abstract B011: Single-molecule protein footprinting with Fiber-seq resolves coordinated chromatin states across regulatory domain

An integrated single-molecule view of regulatory remodeling may support identification of adaptive resistance mechanisms, compensatory regulatory programs, pharmacodynamic biomarkers, and candidate synthetic lethal interactions relevant to epigenetic drug development.

Pangenome discovery and characterization of human protein-coding duplicated genes

The pangenome provides unparalleled specificity to understand genetic variation in SD genes allowing us to distinguish functional genes from pseudogenes and highlighting potential gene innovations that arose most recently in human evolution.