2001 — 2003 |
Wijnen, Herman |
F32Activity Code Description: To provide postdoctoral research training to individuals to broaden their scientific background and extend their potential for research in specified health-related areas. |
Output of the Circadian Clock
DESCRIPTION (provided by applicant): Circadian (-24 hour) rhythms that are controlled by circadian clocks are found in the physiology and behavior of many organisms including humans and fruitflies. In mammals the sleep-wake cycle is the most obvious example of a circadian rhythm that can have important effects on health, but the regulation of body temperature, heartbeat, blood pressure, endocrine functions, renal activity, and liver metabolism have also been shown to be under circadian regulation. Circadian clocks generate oscillating patterns of transcription that control both the rhythm of the clocks themselves and the output that they produce. It is the goal of this research project to first test on a genome-wide scale which transcripts in the head of adult fruitflies are regulated by circadian clocks and then identify a subset of these transcripts that are involved in the regulation of the rest-activity cycle. DNA microarrays will be used to perform the expression analyses; locomotor activity assays will serve as a phenotypic measure for the rest-activity cycle; and a combination of genetic and molecular techniques will be employed in the functional analysis of candidate regulators of the rest-activity cycle. The comprehensive description of the clock controlled transcripts in Drosophila heads will provide a valuable resource for all future studies on circadian rhythms, whereas the discovery of new mediators of the rest-activity cycle will promote a better understanding of the molecular mechanisms underlying both the rest-activity cycle in fruitfiles and the analogous sleep-wake cycle in mammals.
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0.946 |
2007 — 2011 |
Wijnen, Herman |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Transcriptional Output Pathways of the Circadian Clock @ University of Virginia Charlottesville
[unreadable] DESCRIPTION (provided by applicant): Circadian clocks in the bodies of humans ensure that functions such as the sleep/wake cycle, body temperature, hormonal secretion, cardiovascular function, respiration and metabolism are timed correctly. There is an exceptionally high degree of conservation between the circadian clocks of mammals and insects and much of our current understanding of clock function in humans is based on studies of the genes and the molecular circuits of the clock in model organisms such as Drosophila. Transcriptional feedback circuits responsible for the autonomous pacemaker function of the Drosophila clock have been identified in detail, but much less is known about the pathways connecting these circuits to overt rhythms in physiology and behavior. This proposal is designed to test the hypothesis that the CLK/CYC transcription factor of the Drosophila clock circuits has direct transcriptional targets in the Drosophila brain that generate behavioral rhythms. Specific Aims: (1) Transcriptional targets specific to the circadian transcription factor CLK/CYC will be identified with the use of a set of transgenic flies that allow expression of each of these transcription factors to be induced individually in clock-bearing cells. (2) Direct transcriptional targets for CLK/CYC will be identified by luciferase reporter assays and chromatin immunoprecipitation experiments performed in Drosophila S2 cells. (3) Locomotor and eclosion behavior phenotypes associated with primary transcriptional clock output in the brain will be determined with the use of transgenic flies. Significance: The insight provided by the proposed studies into the internal time keeping mechanisms of animals is not only relevant to diseases associated with disruptions of these mechanisms (sleep disorders, cancer, diabetes), but also more generally to diagnostic and treatment procedures involving bodily functions that show regular daily variations. [unreadable] [unreadable] [unreadable]
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0.946 |
2009 |
Wijnen, Herman |
S10Activity Code Description: To make available to institutions with a high concentration of NIH extramural research awards, research instruments which will be used on a shared basis. |
Bioluminescence/Fluorescence Imaging System For Long-Term in Vivo Recording
DESCRIPTION (provided by applicant): Many physiological parameters affecting human health and disease, including the sleep/wake cycle, core body temperature, hormonal secretion, cardiovascular function, respiration, and metabolism exhibit daily rhythms that are generated by an internal circadian clock. Studies of the mechanisms producing these rhythms may enable the development of improved protocols for diagnosis and treatment of disease that take circadian variation into account. In addition, circadian clock defects have been directly associated with intrinsic, circadian rhythm sleep disorders, cancer, and metabolic disorders. The circadian clocks of humans and other eukaryotes are gene expression feedback circuits that can be studied directly by monitoring circadian gene expression rhythms. The use of circadian luciferase reporter genes in combination with extremely sensitive bioluminescence detection has enabled high frequency monitoring of the molecular clock function in living cells and tissues. Moreover, the recent development of systems capable of imaging the very weak, but reliable luciferase-mediated bioluminescence has allowed high frequency and high resolution analysis of molecular clock function in individual cells and across cellular networks. The proposed 3D bioluminescence/fluorescence imaging system will be used to image circadian luciferase reporters in cultured insect and mammalian cells and tissues in reference to co-expressed fluorescent markers associated with specific cell types or biological activities. These studies will focus on the cell-autonomous and network properties of circadian clock function in Drosophila and mammals. 3D bioluminescence/fluorescence imaging will be used in combination with the powerful genetic tools and behavioral assays available in Drosophila to dissect the clock-controlled gene expression rhythms that govern behavior and the role that serotonergic pathways play in this context. Mammalian clock function will be imaged in the Gonadotrophin-releasing hormone neurons and Pars Tuberalis of the brain as well as the ovaries and tested for responses to different photoperiods, hormones, neuropeptides and/or neurotransmitters. In addition, mammalian retinas will be imaged to verify the presence of circadian clocks in dopaminergic amacrine cells and test if the formation of gap junctions at intrinsically photosensitive retinal ganglion cells is clock-gated. Finally, the proposed instruments will help delineate the function of the mammalian core clock components CRYPTOCHROME 1 and 2 by allowing both clock-controlled luciferase expression and the expression level and subcellular localization of fluorescently tagged transfected CRY to be monitored in parallel cultured cells. PUBLIC HEALTH RELEVANCE: The proposed instrumentation and experiments will provide insight in the internal daily time keeping mechanisms of humans and animals. Results from the proposed studies will not only help clarify why and how the human body shows functional changes with daily time, but also shed light on the basis for diseases linked to time keeping defects.
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0.946 |
2009 — 2012 |
Hirsh, Jay (co-PI) [⬀] Menaker, Michael (co-PI) [⬀] Provencio, Ignacio (co-PI) [⬀] Green, Carla (co-PI) [⬀] Wijnen, Herman |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Mri: Acquisition of a Bioluminescence/Fluorescence Imaging System For Long-Term Recording of Living Tissues @ University of Virginia Main Campus
This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).
With this award to the Biology department at the University of Virginia a bioluminescence/biofluorescence imaging system will be acquired to study molecular signals associated with daily time keeping mechanisms. The instrument will be used for experiments aimed at gaining a better understanding of the internal circadian clocks that allow animals to anticipate daily changes in their environment and to organize a multitude of bodily functions in an optimized daily schedule. The new imaging system optimally visualizes bioluminescent signals produced by clock-controlled expression of luciferase reporter genes in combination with fluorescent signals produced by additional reporter genes that act as markers for specific cell types or biological activities. With the new technology it will be possible to image cell-type specific circadian gene expression at single cell resolution. The proposed experiments will focus on the cellular and network properties of circadian clock function in fruit fly and rodent model systems. These research activities will strongly stimulate the research programs of the (Co-)PIs as well as help in the recruitment and training of researchers and students to the University of Virginia. The instrument and planned research activities will be prominently featured in several undergraduate courses. In addition, existing affiliations will be used to also incorporate training and outreach at the K-12 level. Participating researchers and students will not only be involved in projects at the cutting edge of chronobiological research and imaging technology, but also receive invaluable training in microscopy and data analysis techniques that will be an asset to them throughout their scientific or professional careers.
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0.943 |