Revealing the molecular mechanisms of the circadian clock
Nobel Prizes in Physiology or Medicine announced
Life on Earth is adapted to the rotation of our planet. For many years we have known that living organisms, including humans, have an internal, biological clock that helps them anticipate and adapt to the regular rhythm of the day. But how does this clock actually work? Jeffrey C. Hall, Michael Rosbash and Michael W. Young were able to peek inside our biological clock and elucidate its inner workings. Their discoveries explain how plants, animals and humans adapt their biological rhythm so that it is synchronized with the Earth's revolutions.
Figure 1. An internal biological clock. The leaves of the mimosa plant open towards the sun during day but close at dusk (upper part). Jean Jacques d'Ortous de Mairan placed the plant in constant darkness (lower part) and found that the leaves continue to follow their normal daily rhythm, even without any fluctuations in daily light.
© The Nobel Committee for Physiology or Medicine. Illustrator: Mattias Karlén
Figure 2A. A simplified illustration of the feedback regulation of the period gene. The figure shows the sequence of events during a 24h oscillation. When the period gene is active, period mRNA is made. The mRNA is transported to the cell's cytoplasm and serves as template for the production of PER protein. The PER protein accumulates in the cell's nucleus, where the period gene activity is blocked. This gives rise to the inhibitory feedback mechanism that underlies a circadian rhythm.
© The Nobel Committee for Physiology or Medicine. Illustrator: Mattias Karlén
Figure 2B. A simplified illustration of the molecular components of the circadian clock.
© The Nobel Committee for Physiology or Medicine. Illustrator: Mattias Karlén
Figure 3. The circadian clock anticipates and adapts our physiology to the different phases of the day. Our biological clock helps to regulate sleep patterns, feeding behavior, hormone release, blood pressure, and body temperature.
© The Nobel Committee for Physiology or Medicine. Illustrator: Mattias Karlén
Using fruit flies as a model organism, this year's Nobel laureates isolated a gene that controls the normal daily biological rhythm. They showed that this gene encodes a protein that accumulates in the cell during the night, and is then degraded during the day. Subsequently, they identified additional protein components of this machinery, exposing the mechanism governing the self-sustaining clockwork inside the cell. We now recognize that biological clocks function by the same principles in cells of other multicellular organisms, including humans.
With exquisite precision, our inner clock adapts our physiology to the dramatically different phases of the day. The clock regulates critical functions such as behavior, hormone levels, sleep, body temperature and metabolism. Our wellbeing is affected when there is a temporary mismatch between our external environment and this internal biological clock, for example when we travel across several time zones and experience "jet lag". There are also indications that chronic misalignment between our lifestyle and the rhythm dictated by our inner timekeeper is associated with increased risk for various diseases.
Our inner clock
Most living organisms anticipate and adapt to daily changes in the environment. During the 18th century, the astronomer Jean Jacques d'Ortous de Mairan studied mimosa plants, and found that the leaves opened towards the sun during daytime and closed at dusk. He wondered what would happen if the plant was placed in constant darkness. He found that independent of daily sunlight the leaves continued to follow their normal daily oscillation (Figure 1). Plants seemed to have their own biological clock.
Other researchers found that not only plants, but also animals and humans, have a biological clock that helps to prepare our physiology for the fluctuations of the day. This regular adaptation is referred to as the circadian rhythm, originating from the Latin words circa meaning "around" and dies meaning "day". But just how our internal circadian biological clock worked remained a mystery.
Identification of a clock gene
During the 1970's, Seymour Benzer and his student Ronald Konopka asked whether it would be possible to identify genes that control the circadian rhythm in fruit flies. They demonstrated that mutations in an unknown gene disrupted the circadian clock of flies. They named this gene period. But how could this gene influence the circadian rhythm?
This year's Nobel Laureates, who were also studying fruit flies, aimed to discover how the clock actually works. In 1984, Jeffrey Hall and Michael Rosbash, working in close collaboration at Brandeis University in Boston, and Michael Young at the Rockefeller University in New York, succeeded in isolating the period gene. Jeffrey Hall and Michael Rosbash then went on to discover that PER, the protein encoded by period, accumulated during the night and was degraded during the day. Thus, PER protein levels oscillate over a 24-hour cycle, in synchrony with the circadian rhythm.
A self-regulating clockwork mechanism
The next key goal was to understand how such circadian oscillations could be generated and sustained. Jeffrey Hall and Michael Rosbash hypothesized that the PER protein blocked the activity of the period gene. They reasoned that by an inhibitory feedback loop, PER protein could prevent its own synthesis and thereby regulate its own level in a continuous, cyclic rhythm (Figure 2A).
The model was tantalizing, but a few pieces of the puzzle were missing. To block the activity of the period gene, PER protein, which is produced in the cytoplasm, would have to reach the cell nucleus, where the genetic material is located. Jeffrey Hall and Michael Rosbash had shown that PER protein builds up in the nucleus during night, but how did it get there? In 1994 Michael Young discovered a second clock gene, timeless, encoding the TIM protein that was required for a normal circadian rhythm. In elegant work, he showed that when TIM bound to PER, the two proteins were able to enter the cell nucleus where they blocked period gene activity to close the inhibitory feedback loop (Figure 2B).
Such a regulatory feedback mechanism explained how this oscillation of cellular protein levels emerged, but questions lingered. What controlled the frequency of the oscillations? Michael Young identified yet another gene, doubletime, encoding the DBT protein that delayed the accumulation of the PER protein. This provided insight into how an oscillation is adjusted to more closely match a 24-hour cycle.
The paradigm-shifting discoveries by the laureates established key mechanistic principles for the biological clock. During the following years other molecular components of the clockwork mechanism were elucidated, explaining its stability and function. For example, this year's laureates identified additional proteins required for the activation of the period gene, as well as for the mechanism by which light can synchronize the clock.
Keeping time on our human physiology
The biological clock is involved in many aspects of our complex physiology. We now know that all multicellular organisms, including humans, utilize a similar mechanism to control circadian rhythms. A large proportion of our genes are regulated by the biological clock and, consequently, a carefully calibrated circadian rhythm adapts our physiology to the different phases of the day (Figure 3). Since the seminal discoveries by the three laureates, circadian biology has developed into a vast and highly dynamic research field, with implications for our health and wellbeing.
Original publication
Zehring, W.A., Wheeler, D.A., Reddy, P., Konopka, R.J., Kyriacou, C.P., Rosbash, M., and Hall, J.C.; "P-element transformation with period locus DNA restores rhythmicity to mutant, arrhythmic Drosophila melanogaster"; Cell; 1984
Bargiello, T.A., Jackson, F.R., and Young, M.W.; "Restoration of circadian behavioural rhythms by gene transfer in Drosophila"; Nature; 1984
Siwicki, K.K., Eastman, C., Petersen, G., Rosbash, M., and Hall, J.C.; "Antibodies to the period gene product of Drosophila reveal diverse tissue distribution and rhythmic changes in the visual system"; Neuron; 1988
Hardin, P.E., Hall, J.C., and Rosbash, M.; "Feedback of the Drosophila period gene product on circadian cycling of its messenger RNA levels"; Nature; 1990
Liu, X., Zwiebel, L.J., Hinton, D., Benzer, S., Hall, J.C., and Rosbash, M.; "The period gene encodes a predominantly nuclear protein in adult Drosophila"; J Neurosci; 1992
Vosshall, L.B., Price, J.L., Sehgal, A., Saez, L., and Young, M.W.; "Block in nuclear localization of period protein by a second clock mutation, timeless"; Science; 1994
Price, J.L., Blau, J., Rothenfluh, A., Abodeely, M., Kloss, B., and Young, M.W.; "double-time is a novel Drosophila clock gene that regulates PERIOD protein accumulation"; Cell; 1998
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