Abstract
Through the use of bulk measurements in metabolic organs, the circadian clock was shown to play roles in organismal energy homeostasis. However, the relationship between metabolic and circadian oscillations has not been studied invivo at a single-cell level. Also, it is unknown whether the circadian clock controls metabolism in stem cells. We used a sensitive, noninvasive method to detect metabolic oscillations and circadian phase within epidermal stem cells in live mice at the single-cell level. We observe a higher NADH/NAD+ ratio, reflecting an increased glycolysis/oxidative phosphorylation ratio during the night compared to the day. Furthermore, we demonstrate that single-cell metabolic heterogeneity within the basal cell layer correlates with the circadian clock and that diurnal fluctuations in NADH/NAD+ ratio are Bmal1 dependent. Our data show that, in proliferating stem cells, the circadian clock coordinates activities of oxidative phosphorylation and glycolysis with DNA synthesis, perhaps as a protective mechanism against genotoxicity.
Highlights
The circadian clock is a self-sustained cellular oscillator that coordinates appropriate metabolic responses within peripheral tissues with the light/dark cycle
We first performed in vivo imaging of NADH autofluorescence and collagen second harmonic generation (SHG) in adult mouse skin (Figures 1A–1C) to determine whether we could utilize the SHG from the dermis to localize cells of the basal cell layer
Analysis of the fluorescence lifetime imaging microscopy (FLIM) images is performed by a fast Fourier transform (FFT) of the FLIM raw data (Figures 1D and 1E) by creating a 2D histogram of the NADH FLIM image where every pixel of the FLIM image is transformed into a pixel in the phasor plot (Figures 2A and 2B)
Summary
The circadian clock is a self-sustained cellular oscillator that coordinates appropriate metabolic responses within peripheral tissues with the light/dark cycle. In vivo evaluations of metabolic oscillations have been done through bulk-tissue experiments It remains unknown whether the circadian clock is involved in metabolism control in stem cells that maintain self-renewing epithelia. More recent studies have started to cast light on how the time-of-day-dependent variation in cell proliferation is regulated, showing that core circadian clock components are required for this feature (Gaddameedhi et al, 2011; Geyfman et al, 2012; Janich et al, 2011, 2013; Plikus et al, 2013)
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