Single-molecule chromatin tracing reveals a diversity of megabase heterochromatin domains

Abstract Chromosomes fold into distinct domains that regulate transcription, replication and repair. In addition to well-characterized topologically associating domains (TADs) and compartments, the diversity of heterochromatin domains remains poorly defined at the sequence level. Here, using single-molecule tracing of nascent heterochromatin in Caenorhabditis elegans , we identify three classes of megabase-scale domains: (1) sharp-boundary, condensin I-dependent TAD-like domains (TADLs), which arise before zygotic genome activation; (2) similarly sized, but condensin-independent, el egans condensin-independent domains (elCIDs), one of which encompasses the 5S-spliced leader repeat locus; and (3) decompacted, diffuse structures that are abundant in the population but probably missed by crosslinking assays. Condensin I sits at the border of the TADL domain, whereas DNA-binding proteins, active histone marks and maternally active genes flank the elCID domains, suggesting that boundaries are established by distinct mechanisms for different types of domains. Our results uncover previously unrecognized heterochromatin architectures and implicate maternal transcriptional activity in shaping domain boundaries in early embryos in C.   elegans .