STEM CELL O’ CLOCK: CIRCADIAN TIMING AS A REGULATOR OF ADULT NEURAL STEM CELL DIVISION AND MOLECULAR FATE DYNAMICS

Adult NSCs in the dentate gyrus are predominantly quiescent, with activation tightly regulated to balance neurogenesis and stem cell maintenance. Circadian clocks temporally organize core cellular processes. Previous evidence suggests that the circadian clock influences adult neural stem cell proliferation. Here, we further examined its functionality, impact in coordinated activation, and the underlying circadian transcriptional programs.
We examined NSCs proliferation dynamics across the day in PER2::VENUS reporter mice and assessed whether NSCs possess functional cellular circadian clocks. We observed rhythmic expression of core clock components in NSCs (BMAL1 and PER2) together with a nighttime increase in S-phase cells (EdU⁺ NSCs). This coordinated activation was abolished in Per2 knockout mice, indicating that PER2 is required for circadian gating of adult NSCs.
To determine whether NSCs exhibit intrinsic circadian rhythmicity, we performed time-resolved transcriptomic analysis of NSCs in vitro, sampling every four hours over 48 hours. Active NSCs showed greater rhythmic transcription, with clock rhythmicity in both states but a shift from negative-loop transcripts (Per2, Cry1, Cry2, Bhlhe41) in active cells to positive-loop transcripts (Bmal1, Clock) in quiescence. Immunocytochemistry further validated core clock activity at a protein level (BMAL1 and PER2).
Comparative analysis of WT and Per2KO NSCs revealed differential state-specific, clock-dependent oscillatory programs in quiescent cells and active NSCs. Specific candidates identified in circadian transcriptomics were subsequently confirmed by antibody stainings in vivo. Together, these findings reveal state-specific circadian regulation of adult NSCs with implications for temporal control of activation and fate decisions.