SMCHD1 loss re-wires MYOD1 enhancer nexuses and chromatin accessibility landscapes in muscle cells

Human SMCHD1 (Structural Maintenance of Chromosomes Flexible Hinge Domain Containing 1) is a chromatin architectural protein linked to heterochromatin repression. Loss of function mutations of SMCHD1 cause facioscapulohumeral muscular dystrophy type 2 (FSHD2) through activation of the <i>DUX4</i> homeobox transcription factor gene. However, it is unknown how SMCHD1 may regulate myogenic transcription independently of DUX4. Here, we show that SMCHD1 safeguards enhancer organization within the three-dimensional (3D) genome in human myoblasts. Loss of SMCHD1 leads to widespread gains in chromatin accessibility, aberrant transcription and a global redistribution of the myogenic transcription factor MYOD1. Integrative analyses of histone modifications, chromatin accessibility, Hi-C looping, and activity-by-contact enhancer-gene modeling reveal that SMCHD1 loss rewires the landscape of clustered enhancers and promotes the emergence of a new MYOD1-related network of enhancer elements, termed MYOD1 enhancer nexuses. These structures are marked by increased enhancer-enhancer connectivity, increased local 3D chromatin interactions, and coordinated activation of genes likely relevant for FSHD pathology. Together, our findings identify SMCHD1 as a key architectural constraint that suppresses hyperactive enhancer networks, thereby preserving transcriptional homeostasis in myoblasts.