The c-MYC proto-oncogene regulates cellular proliferation, and its aberrant expression drives a range of human cancers. It also has a bidirectional regulatory relationship with the mammalian core circadian clock, with emerging evidence suggesting that MYC overexpression leads to clock disruption and loss of rhythms. While prior studies have probed the role of MYC role in clock disruption by overexpressing or mutating the c-MYC gene, our understanding of the endogenous nature of c-MYC is limited. A major gap in knowledge is whether MYC itself is expressed rhythmically and if so, how its timing relates to that of core clock components. To address these shortcomings, we generated a c-MYC reporter and assessed its circadian nature, comparing it to BMAL1 and PER2, and developed a computational model based on these and previous findings to evaluate its role(s). We developed lentiviral constructs for and established a U2OS (common circadian model) reporter cell line expressing luciferase (luc) driven by a human-derived c-MYC promoter sequence. To facilitate comparisons, as part of this work, we also developed a human-sequence derived BMAL1 promoter reporter to more readily recapitulate its behaviors. Using luminometry studies and subsequent data analyses, we demonstrated that the c-MYC promoter oscillated rhythmically in U2OS cells, which possess inherently low levels of c-MYC. Furthermore, we found that c-MYC oscillates out-of-phase relative to BMAL1 and PER2. Using this information, we built a mathematical model to better understand how c-MYC oscillations at both basal and over-expressed levels affect the clock and vice versa. The model reproduced expected alterations to the core clock resulting from c-MYC overexpression and showed that MYC acts as a disruptor, although the timing of MYC regulation can minimize its negative impact(s) on circadian timekeeping. This work is the first to assess the phase relationships of c-MYC relative to the core clock and to provide evidence for its circadian nature.
Kalyanaraman, B., Ganesh, D., Kunte, V. A., Taylor, S. R., Farkas, M. E.
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