The molecular basis of the mammalian circadian clock
Giles E. Duffield
Assistant Professor
Ph.D., University of Cambridge, 1998
Postdoctoral and Faculty Research (Wellcome Trust Research Fellow, Royal Society University Research Fellow), Dartmouth Medical School and Imperial College London |
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New Postdoctoral Research Associate Position Available
Circadian ('about a day') rhythms are an integral component of biochemistry, physiology and behavior. Circadian clock biology is relevant to human health. Dysfunction of the circadian clock underlies several disease states, including Seasonal Affective Disorder, and sleep disorders. My lab is interested in elucidating the molecular basis of the circadian clock in mammals using a range of traditional and state of the art molecular, cellular and behavioral approaches.
The molecular circadian clock consists of an autoregulatory transcriptional-translational feedback loop composed of positive and negative regulators. Work over the last 8 years has identified 9 such components, but additional genes and modifiers are being identified. In addition, most tissues of the body harbor cell-autonomous circadian clocks.
Alan J. Hesse, Cartoons for Conservation
One such additional gene, identified in my laboratory using a cDNA microarray screen, is the transcriptional inhibitor Inhibitor of DNA-binding 2. It is rhythmically expressed in the master clock structure in the hypothalamic brain known as the suprachiasmatic nucleus (SCN), and throughout the body in various peripheral tissues (e.g. heart and liver). Current studies are to evaluate the role of genes such as Id2 in the organization of the central oscillator, and to identify novel molecules relevant to cellular clock function. We are also interested in understanding how light resets the molecular clock (input), and how the clock regulates down-stream clock-controlled genes (i.e. output, hands of the clock).
My lab is using continuous activity monitoring to identify behavioral phenotypes in transgenic mouse models (e.g. Id2 knockout mice) that are maintained under a variety of photocycle conditions, and exposed to artificial time-zone changes and acute light/pharmacologic/behavioral treatments. Results thus far have revealed that in the absence of the Id2 gene, mice adapt to large time-zone changes (e.g. mimicking a flight from Berlin to Los Angeles) more rapidly than wildtype individuals. We are also using real-time monitoring of clock gene expression in tissues and cells derived from transgenic mice that express Firefly luciferase in a rhythmic manner. We are using DNA microarray and real-time quantitative RT-PCR analyses to identify and characterize clock regulated genes, tissue culture of immortalized fibroblasts that exhibit circadian oscillations in gene expression as a model of the in vivo rodent circadian clock, and traditional neuroanatomical techniques (e.g. in situ hybridization, immunohistochemistry, neuronal track tracing) to characterize clock gene function in the brain. These studies have been supported by the Royal Society, the Wellcome Trust and the NIMH.
Northern blot analysis of mouse NIH3T3 fibroblasts, reveals rhythmic gene expression of canonical clock components
cDNA microarray analysis of mouse NIH3T3 fibroblasts to search for rhythmic clock controlled genes
Immunohistochemical staining of the developing suprachiasmatic nuclei
Research Personnel
Maricela Robles Murquia, Research Technician
Sam Rund, Graduate Student
Sarah Ward, Graduate Student
Tim Hou, Research Associate
Kathleen McDonald, undergraduate student
Kevin Flanagan, undergraduate student
Shanik Fernando, undergraduate student
Collaborators
• Mark Israel, Director, Norris Cotton Cancer Center, Dartmouth Hitchcock Medical Center
• Jay Dunlap and Jennifer Loros, Department of Genetics, Dartmouth Medical School
• Horacio de la Iglesia, Department of Biology, University of Washington
• Stuart Peirson, Senior Research Scientist, Department of Ophthalmology, University of Oxford
• Nelson Chong, Lecturer in Molecular Cardiology, Department of Cardiovascular Sciences, Leicester Medical School, University of Leicester
Selected Bibliography
Hou, T., Ward, S.M., Murad J.M., Watson, N.P., Israel, M.A. and Duffield, G.E. (2009) Inhibitor of DNA binding 2 (ID2) is a rhythmically expressed transcriptional repressor required for circadian clock output in the mouse liver. Journal of Biological Chemistry 2009 Nov 13; 284:31735-45 [Epub 2009 Sep 9].
Duffield G.E., Robles-Murguia M., Watson N.P., Mantani A., Peirson S.N., Loros J.J., Israel M.A., Dunlap J.C. (2009) A role for the Id2 gene in regulating photic entrainment of the mammalian circadian system. Current Biology; 19:297-304.
Peirson, S.N., Butler, J.N., Duffield, G.E., Takher, S., Sharma, P. and Foster, R.G. (2006). Comparison of clock gene expression in SCN, retina, heart and liver of mice. Biochemical and Biophysical Research Communications; 351:800-807.
Duffield, G., Loros, J.J., and Dunlap, J.D. (2005) Analysis of circadian output rhythms of gene expression in neurospora and mammalian cells in culture. Methods Enzymology 393:315-341. Invited paper for edition on Circadian Rhythms.
Duffield, G.E. (2003) DNA microarray analyses of circadian timing (Invited Review, 'Young Investigator's Perspectives'). Journal of Neuroendocrinology 15:991-1002.
Nowrousian, M., Duffield, G.E., Loros, J.J. and Dunlap, J.C. (2003) The frequency gene is required for temperature-dependent regulation of many clock-controlled genes in Neurospora crassa. Genetics 164:923-933.
Duffield, G.E., Best, J.D., Meurers, B.H., Bittner, A., Loros, J.J. and Dunlap, J.C. (2002) Circadian programs of transcriptional activation, signaling, and protein turnover revealed by microarray analysis of mammalian cells. Current Biology 12:551-557, April 3 (cover article).
Duffield, G.E., McNulty, S. and Ebling, F.J.P. (1999) Anatomical and funcational characterization of a dopaminergic system in the suprachiasmatic nucleus of the neonatal Siberian hamster. Journal of Comparative Neurology 408:73-96.
von Gall, C., Duffield, G.E., Hastings, M.H., Kopp, M.D.A., Dehghani, F., Korf, H.-W., Stehle, J.H. (1998). CREB in the Mouse SCN: A Molecular Interface Coding the Phase-Adjusting Stimuli Light, Glutamate, PACAP, and Melatonin for Clockwork Access. Journal of Neuroscience 18: 10389-10397.
Duffield G.E., Hastings M.H. and Ebling F.J.P. (1998). Investigation into the regulation of the circadian system by dopamine and melatonin in the Siberian hamster (Phodopus sungorus). Journal of Neuroendocrinology 10: 871-884.
Department of Biological Sciences
Galvin Life Science Center
Notre Dame, IN 46556-0369
Office Ph: (574) 631-1834
Lab Ph: (574) 631-1869
Fax: (574) 631-7413
E-mail: Giles.E.Duffield.2@nd.edu
