We all suffer from jet lag when we fly long distances east or west and this is because our internal clock and our wristwatch are not telling us the same time. The internal circadian clock helps us sense, anticipate and adapt to the periodic changes that happen during the 24 hr day. Sleeping sickness is the common name of an infectious disease caused by Trypanosoma brucei parasite. In this disease, sleep becomes fragmented and people sleep more during the day and have insomnia at night.
When Filipa Rijo-Ferreira decided to do her PhD in my laboratory, her question was whether parasites interfered with the circadian rhythm of the host, which would ultimately lead to the characteristic sleep changes. Joe Takahashi (UT Southwestern), an expert in circadian rhythm in mice, loved this idea but a second immediate question emerged from our conversation: do trypanosomes also have an internal circadian clock? We debated about this. On one hand, parasites, like probably all organisms on Earth, are exposed to diurnal environmental changes and thus it was likely that during evolution, a clock mechanism would have been advantageous for the parasite fitness. On the other hand, parasites are never free-living and, moreover, other virulence mechanisms are probably much more important for their fitness. This was a risky project! But given that trypanosomes are extracellular, they represented the ideal model system to test whether non-free living infectious organisms have a mechanism of counting time.
Filipa created a trans-Atlantic "shuttle" between Lisbon and Dallas (Video below). She traveled back and forth doing the parasitology side of this project in Lisbon, and taking advantage of the tools and expertise in circadian rhythms in Dallas. Our approach was to grow parasites in culture, extract their RNA every four hours for two days and perform RNA-seq experiments. We wanted to know whether there were any genes, whose transcript levels oscillated in a periodic manner. Normally, the clocks of individual cells in culture rapidly become asynchronous. For this reason, we needed to synchronize the parasite cultures with an external stimulus, prior to extracting RNA. The bioinformaticians in our team found that, when parasites were synchronized with warm/cold cycles, around 10% of the genes showed a periodic oscillation at the RNA level. Many of these genes encode for proteins involved in metabolism. Next we confirmed that the oscillations of transcript levels were temperature entrained and temperature compensated.
Behind the paper – Trypanosoma brucei metabolism is under circadian control from Fernando Augusto on Vimeo.
T. brucei parasites in the bloodstream divide every 7 hours. The fact that we detected variations in the transcriptome with a 24 hr period indicates that: (i) parasites in the morning are different from those in the afternoon; and, (ii) parasites pass on the "time information" to the daughter cells when they divide. Overall, our data show that ∼10% of the transcriptome undergoes circadian oscillations, suggesting that having a circadian rhythm might have conferred an evolutionary advantage to the parasite. One of the next obvious questions is to determine what this advantage is. The other question is to identify the components of this molecular clock in trypanosomes.
There is a lot to do! When will we be able to sleep?
The paper in Nature Microbiology is here: http://go.nature.com/2mHE4wh
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