Flying half way across the world, staying up late to play on your smart phone and pulling an all-nighter to cram for tomorrow’s exam can all upset your body’s daily rhythms. These rhythms, which are called circadian rhythms, are roughly 24 hour oscillations in behavior and physiology that function to anticipate the environmental changes associated with the solar day. These rhythms are internally generated and driven by a circadian clock that responds to light and darkness from the environment.
Circadian rhythms can influence a variety of things in the body such as sleep-wake cycles, body temperature and hormone release. Abnormal circadian rhythms have been associated with a variety of conditions such as insomnia, diabetes and obesity. Understanding how our clocks work is therefore critical to the development of drugs for these diseases.
While previous research had identified a set of four core clock genes-- Cryptochrome, Period, CLOCK and BMAL1-- that together generate rhythmicity in cells, the precise roles played by these genes and how they interact was unclear. Now, a team of scientists from the University of North Carolina have finally pieced together these individual components and deciphered how the entire clock works, and how it is reset in cells.
Previous work discovered that CLOCK and BMAL1 work in concert to start the circadian clock in cells. The proteins produced by these genes are both transcription factors, which are proteins that bind to specific stretches of DNA in order to control gene expression. It was found that CLOCK and BMAL1 proteins form a complex that binds to the Period and Cryptochrome genes, switching them on and initiating gene expression. The proteins produced by this second set of genes then suppress CLOCK and BMAL1, which in turn represses their own expression. When Period and Cryptochrome proteins are eventually degraded, the clock can restart.
“It’s a feedback loop,” senior author Aziz Sancar said in a news-release. “The inhibition takes 24 hours.”
While this much was known, scientists didn’t know how CLOCK and BMAL1 were suppressed, or what triggered the degradation of Period and Cryptochrome. To find out more, researchers knocked out both the Cryptochrome and Period genes in cells. When they re-added Period into these cells, they found that it could not inhibit the CLOCK:BMAL1 complex. Next, they tried just adding Cryptochrome back into the cells. Cryptochrome alone successfully inhibited CLOCK and BMAL1, but it did so irreversibly because it was not degraded.
Lastly, the researchers tried adding Period to this second set of cells. They found that as the Period protein accumulated, it gradually started to remove not only Cryptochrome but CLOCK and BMAL1 too. This eventually triggered the degradation of Cryptochrome, freeing up the CLOCK and BMAL1 genes to restart the clock and thus completing the 24 hour cycle.
“What we’ve done is show how the entire clock really works,” said Sancar. “Now, when we screen for drugs that target these proteins, we know to expect different outcomes and why we get those outcomes.”
The team’s work has been published in the journal Genes & Development.