The present research will explore and develop a transgenic approach to producing a bioluminescent reporter strain of Syrian hamster that will be a valuable tool for circadian research. The tau hamster was the first mammalian period mutant to be discovered. Transplantation experiments using tau hamsters established that suprachiasmatic nucleus (SCN) regulates the daily scheduling of physiology and behavior. Discovery that tau is a gain of function of casein kinase 1? was critical to achieving an understanding of the operation of transcriptional-translational feedback loops that generate circadian rhythms, both in the brain's pacemaker and in peripheral oscillators. We recently discovered duper, a new mutation in hamsters which, like tau, speeds up the circadian clock. Duper is not a change in the coding region of casein kinase or any other known clock gene, and the mutation does not affect clock speed in fibroblasts. Thus duper is unlikely to affect the TTFL, but more probably alters coupling relationships within the SCN. Duper causes a striking reduction in jet lag. Our experiments will validate the reporter strain by comparing luminescent traces with results from qRT-PCR. We will then compare the impact of the tau and duper mutations on circadian rhythms in order to determine whether shortening of period through effects on the pacemaker vs alterations in the TTFL have different consequences for organismal function. Although hamsters have provided a valuable model for studies of biological rhythms, their potential as a genetic tool has yet to be realized. We have recently contributed to the first draft of the hamster genome and established methods for making transgenic hamsters. We have employed piggyBac methodology to insert circadian reporters into a hamster cell line. This technology will now enable us to study organismal function. In the first application of the reporter strain, we will determine effects of duper upon the core cell-autonomous feedback loops and upon the coordination of pacemaker function. This will shed light on the mechanisms of circadian desynchrony. Unlike the species in which transgenic reporter strains have thus far been produced, hamsters have a particularly regular, circadian-based estrous cycle and a strong photoperiodic response. Thus development of the hamster reporter strain will make possible future applications to investigate neuroendocrine function. Finally, the importance of the hamster as a disease model for several pathologies including MERS and Ebola, and the relevance of jet lag to the incidence of disease in an era of frequent trans-meridian travel, indicate that the reporter strain will be of widespread utility. Given the importance of circadian organization in behavior and physiology, this research will reveal mechanisms that underlie neurologic diseases, sleep deficiencies and metabolic disorders, and lead to development of new therapies.
The proper timing of arousal, sleep, and essential neurological and metabolic processes that are essential to health is governed by circadian oscillators which depend upon intricate and interlocked transcriptional/translational feedback loops whose phase is set by environmental and internal signals. We will develop a transgenic luminescent reporter strain of Syrian hamster that will allow us to track in real time not only the circadian rhythms of wild types, but also those of duper and tau mutant animals in order to increase our understanding of the mechanisms that govern the period and the phase shifting properties of brain and peripheral oscillators. Given that inappropriate or poorly coordinated scheduling of cellular functions increases the risk of disease, and is a risk factor for millions of Americans who engage in shift work or travel across time zones, the basic bench discoveries that result from these studies will identify homeostatic mechanisms and lead to improved therapeutic treatments for neurological disorders and psychiatric illness, as well as cancer and metabolic diseases.
| Manoogian, Emily N C; Kumar, Ajay; Obed, Doha et al. (2018) Suprachiasmatic function in a circadian period mutant: Duper alters light-induced activation of vasoactive intestinal peptide cells and PERIOD1 immunostaining. Eur J Neurosci 48:3319-3334 |
