Nie, X. Y. | bioRxiv Science

bioRxiv2025

Single-Cell Multiomic Analysis of Circadian Rhythmicity in Mouse Liver

Circadian rhythms are remarkably widespread across most organisms, regulating hormonal, metabolic, physiological, and behavioral oscillations through molecular clocks that orchestrate the rhythmic expression of thousands of genes. Here, we generate single-nucleus RNA and ATAC multiomics data to simultaneously characterize gene expression and chromatin accessibility of mouse liver cells across the 24-hour day. We interrogate multimodal circadian rhythmicity in both discretized cell types and transient sub-lobule cell states, capturing space-time omics profiles. We delve beyond mean cyclic patterns to characterize stochastic transcriptional bursting and infer spatiotemporal gene regulatory networks that control circadian rhythmicity and liver physiology. Our findings apply to existing single-cell data of mouse and Drosophila brains and are validated by time-series single-molecule fluorescence in situ hybridization and vast amounts of orthogonal omics data. Altogether, our study constructs a comprehensive map of the time-series transcriptomic and epigenomic landscapes that elucidate the function and mechanism of the liver peripheral clocks.

bioinformatics↗

bioRxiv2024

Circadian regulation of stereotypic chromatin conformations at enhancers

Cooperation between the circadian transcription factor (TF) CLOCK:BMAL1 and other TFs at cis-regulatory elements (CREs) is critical to daily rhythms of transcription. Yet, the mechanisms underlying this cooperation are unclear. Here, we analyzed the co-binding of multiple TFs on single DNA molecules in mouse liver using single molecule footprinting (SMF). We found that SMF reads clustered in stereotypic chromatin states that reflect distinguishable organization of TFs and nucleosomes, and that were remarkably conserved between all samples. DNA protection at CLOCK:BMAL1 binding motif (E-box) varied between CREs, from E-boxes being solely bound by CLOCK:BMAL1 to situations where other TFs competed with CLOCK:BMAL1 for E-box binding. SMF also uncovered CLOCK:BMAL1 cooperative binding at E-boxes separated by 250 bp, which structurally altered the CLOCK:BMAL1-DNA interface. Importantly, we discovered multiple nucleosomes with E-boxes at entry/exit sites that were removed upon CLOCK:BMAL1 DNA binding, thereby promoting the formation of open chromatin states that facilitate DNA binding of other TFs and that were associated with rhythmic transcription. These results demonstrate the utility of SMF for studying how CLOCK:BMAL1 and other TFs regulate stereotypical chromatin states at CREs to promote transcription.

molecular biology↗