Small molecule probes of clock biology and disease
Biological clocks exert temporal control over essential cellular and physiological processes in diverse organisms. Conversely, dysregulated clocks have been associated with a number of chronic diseases and aging. We previously showed that impaired cell cycle gating by the metabolic cycle in yeast results in highly elevated mutation rate, and a conserved DNA damage response pathway couples the cell cycle to the metabolic and circadian cycles (Chen et al., 2007a and 2007b). More recently, we switched focus to the mammalian circadian clock, and conducted a high-throughput chemical screen which led to identification of diverse small molecules capable of modulating the circadian period, phase, amplitude, and damping rate in cells and tissue explants (Fig. 1; Chen et al., 2012). The current focus of my lab is to exploit clock-modulating small molecules as chemical probes of normal and aberrant clocks in respective physiological and pathological settings.
1. Function of the clock-modulating small molecules
We are currently investigating the metabolic and behavioral functions of these molecules in mice. Given the close association between aberrant clocks and various chronic diseases, we are also eager to exploit clock pathways as potential drug targets. Stable cell lines and animal disease models are utilized to determine the functional outcome of manipulating aberrant clocks with small molecules, particularly a subset of clock-enhancing molecules.
2. Mechanism and target of the small molecules
We are systematically analyzing the effect of the small molecules on the molecular oscillator. Furthermore, we wish to understand the cellular signaling pathways underlying their physiological functions, with particular emphasis on metabolic regulation. To identify direct protein targets, we are developing biotinylated derivatives of our very best hits for biochemical affinity pull-down.
3. Second-generation high-throughput screens to identify compounds of novel activities
Taking advantage of the screening facility at the Gulf Coast Consortium located in the Medical School, we are conducting second-generation screens aiming for compounds with novel functions in clock-related cellular processes. We expect to add new probes to our platform of function and mechanism studies.
Dose responses of compounds that specifically affect the period, phase and/or amplitude of the mammalian circadian clock. Blue: vehicle control; purple, green and red: compounds at 0.5, 1.5 and 5 uM, respectively. Y-axis values represent reporter luciferase reading.
Chen, Z.*, Yoo, S.H., Park, Y.S., Kim, K.H., Wei, S., Buhr, E., Ye, Z.Y., Pan, H.L., and Takahashi, J.S.* (2012). Identification of Diverse Modulators of Central and Peripheral Circadian Clocks by High Throughput Chemical Screening. Proc Natl Acad Sci USA 109, 101-106. (*: Corresponding authors)
Chen, Z., Tu, B.P., and McKnight, S.L. (2009). Biological and medical relevance of circadian and metabolic cycles. Seasonal Affective Disorder, 2nd ed., 43-69. Oxford University Press. Eds. T. Partonen and S. R. Pandi-Perumal.
Bertolucci, C., Colognesi, I., Caruso, P., Aguzzi, J., Chen, Z., Cavallari, N., Foa, A., Tosini, G., Bernardi, F., and Pinotti, M. (2008). Evidence for an overlapping role of CLOCK and NPAS2 transcription factors in liver circadian oscillators. Mol. Cell Biol. 28, 3070-3075.
Chen, Z., Odstrcil, E.A., Tu, B.P., and McKnight, S.L. (2007a). Restriction of nuclear DNA replication to the reductive phase of the metabolic cycle protects genome integrity. Science 316, 1916-1919.
Chen, Z., McKnight, S.L. (2007b). A conserved DNA damage response pathway responsible for coupling the cell division cycle to the circadian and metabolic cycles. Cell Cycle 6, 2906-2912.
Chen, Z. and Manley, J.L. (2003). Core promoter elements and TAFs contribute to the diversity of transcriptional activation in vertebrates. Mol. Cell Biol. 23, 7350-7362.
Chen, Z. and Manley, J.L. (2003). In vivo analysis of the histone 3-like TAF9 and a TAF9-related factor, TAF9L. J. Biol. Chem. 278, 35172-35183.
Chen, Z. and Manley, J.L. (2000). Robust mRNA transcription in chicken DT40 cells depleted of TAF31 suggests both functional degeneracy and evolutionary divergence. Mol. Cell Biol. 20, 5064-5076.