The circadian system may regulate adult neurogenesis via intracellular molecular clock mechanisms or by modifying the environment of neurogenic niches, with daily variation in growth factors or nutrients depending on the animal's diurnal or nocturnal lifestyle. In a diurnal vertebrate, zebrafish, we studied circadian distribution of immunohistochemical markers of the cell division cycle (CDC) in 5 of the 16 neurogenic niches of adult brain, the dorsal telencephalon, habenula, preoptic area, hypothalamus, and cerebellum. We find that common to all niches is the morning initiation of G1/S transition and daytime S-phase progression, overnight increase in G2/M, and cycle completion by late night. This is supported by the timing of gene expression for critical cell cycle regulators cyclins D, A2, and B2 and cyclin-dependent kinase inhibitor p20 in brain tissue. The early-night peak in p20, limiting G1/S transition, and its phase angle with the expression of core clock genes, Clock1 and Per1, are preserved in constant darkness, suggesting intrinsic circadian patterns of cell cycle progression. The statistical modeling of CDC kinetics reveals the significant circadian variation in cell proliferation rates across all of the examined niches, but interniche differences in the magnitude of circadian variation in CDC, S-phase length, phase angle of entrainment to light or clock, and its dispersion. We conclude that, in neurogenic niches of an adult diurnal vertebrate, the circadian modulation of cell cycle progression involves both systemic and niche-specific factors.SIGNIFICANCE STATEMENT This study establishes that in neurogenic niches of an adult diurnal vertebrate, the cell cycle progression displays a robust circadian pattern. Common to neurogenic niches located in diverse brain regions is daytime progression of DNA replication and nighttime mitosis, suggesting systemic regulation. Differences between neurogenic niches in the phase and degree of S-phase entrainment to the clock suggest additional roles for niche-specific regulatory mechanisms. Understanding the circadian regulation of adult neurogenesis can help optimize the timing of therapeutic approaches in patients with brain traumas or neurodegenerative disorders and preserve neural stem cells during cytostatic cancer therapies.

P27Kip1 (CDKN1B) regulates cellular proliferation and senescence, and p27Kip1 deficiency in cancer is strongly correlated with poor prognosis of multiple cancer types. Understanding the mechanism of p27Kip1 loss in cancer and the consequences of restoring p27Kip1 levels is therefore critical for effective management during therapy. Here, SIRT1, a class III histone deacetylase (HDAC), is identified as an important regulator of p27Kip1 expression. Mechanistically, SIRT1 reduces p27Kip1 expression by decreasing p27Kip1 protein stability through the ubiquitin-proteasome pathway. In addition, SIRT1 silencing suppresses NSCLC proliferation and induces senescence in a p27Kip1-dependent manner. Furthermore, SIRT1 silencing dramatically suppresses tumor formation and proliferation in two distinct NSCLC xenograft mouse models. Collectively, these data not only demonstrate that SIRT1 is an important regulator of p27Kip1 but that SIRT inhibition induces senescence and anti-growth potential in lung cancer in vivo. Implications SIRT1 is a key regulator of p27 protein levels and SIRT1 inhibition is a viable strategy for NSCLC therapy by means of p27 reactivation.

Chronic high caloric intake (HCI) is a risk factor for multiple major human disorders, from diabetes to neurodegeneration. Mounting evidence suggests a significant contribution of circadian misalignment and sleep alterations to this phenomenon. An inverse temporal relationship between sleep, activity, food intake, and clock mechanisms in nocturnal and diurnal animals suggests that a search for effective therapeutic approaches can benefit from the use of diurnal animal models. Here, we show that, similar to normal aging, HCI leads to the reduction in daily amplitude of expression for core clock genes, a decline in sleep duration, an increase in scoliosis, and anxiety-like behavior. A remarkable decline in adult neurogenesis in 1-year old HCI animals, amounting to only 21% of that in age-matched Control, exceeds age-dependent decline observed in normal 3-year old zebrafish. This is associated with misalignment or reduced amplitude of daily patterns for principal cell cycle regulators, cyclins A and B, and p20, in brain tissue. Together, these data establish HCI in zebrafish as a model for metabolically induced premature aging of sleep, circadian functions, and adult neurogenesis, allowing for a high throughput approach to mechanistic studies and drug trials in a diurnal vertebrate.

Neurogenesis in the adult brain, a powerful mechanism for neuronal plasticity and brain repair, is altered by aging and pathological conditions, including metabolic disorders. The search for mechanisms and therapeutic solutions to alter neurogenesis requires understanding of cell kinetics within neurogenic niches using a high-throughput quantitative approach. The challenge is in the dynamic nature of the process and multiple cell types involved, each having several potential modes of division or cell fate. Here we show that cell kinetics can be revealed through a combination of the BrdU/EdU pulse-chase, based on the circadian pattern of DNA replication, and a differential equations model that describes time-dependent cell densities. The model is validated through the analysis of cell kinetics in the cerebellar neurogenic niche of normal young adult male zebrafish, with cells quantified in 2D (sections), and with neuronal fate and reactivation of stem cells confirmed in 3D whole-brain images (CLARITY). We then reveal complex alterations in cell kinetics associated with accelerated aging due to chronic high caloric intake. Low activity of neuronal stem cells in this condition persists 2 months after reverting to normal diet, and is accompanied by overproduction of transient amplifying cells, their accelerated cell death, and slow migration of postmitotic progeny. This combined experimental and mathematical approach should allow for relatively high-throughput analysis of early signs of pathological and age-related changes in neurogenesis, evaluation of specific therapeutic targets, and drug efficacy. SIGNIFICANCE STATEMENT Understanding normal cell kinetics of adult neurogenesis and the type of cells affected by a pathological process is needed to develop effective prophylactic and therapeutic measures directed at specific cell targets. Complex time-dependent mechanisms involved in the kinetics of multiple cell types require a combination of experimental and mathematical modeling approaches. This study demonstrates such a combined approach by comparing normal neurogenesis with that altered by diet-induced accelerated aging in adult zebrafish.