In vivo BMAL1 occupancy mapping using MACS-Calling Cards reveals disease-associated retargeting in Cln3 ^Δex7/8...

Astrocytic homeostatic programs, many of which are regulated by the circadian clock, are disrupted early in neurodegenerative disease. The core clock transcription factor (TF) BMAL1 is required for normal astrocyte function, but its role during disease remains unclear. This is partly because methods for identifying cell type-specific TF binding sites are limited. Here, we developed MACS-Calling Cards (MACS-CC), a strategy for mapping astrocyte-specific TF occupancy in vivo. We used MACS-CC to define BMAL1 binding in the Cln3 ^Δex7/8 mouse model of CLN3 disease, a fatal neurodegenerative disorder marked by early astrocyte dysfunction and circadian disruption. BMAL1 binding was extensively redistributed in Cln3 ^Δex7/8 astrocytes: wild-type-specific binding sites enriched near glial differentiation genes, whereas Cln3 ^Δex7/8 -specific sites lacked functional enrichment. Consistent with these changes, Cln3 ^Δex7/8 astrocytes decreased expression of mature astrocyte markers. To define mechanisms underlying BMAL1 retargeting, we tested whether altered chromatin accessibility explained the changes in BMAL1 binding. Although chromatin accessibility was broadly remodeled, differential accessibility did not predict BMAL1 redistribution. Instead, motif analysis suggested that loss of cooperative TF interactions drives BMAL1 retargeting. These findings demonstrate that MACS-CC enables astrocyte-specific TF occupancy mapping and reveals mechanisms behind early rewiring of circadian regulatory programs within a model of a neurodegenerative disease.